text:
What are the common methods to estimate additive genetic variance?
Additive genetic variance can be estimated in a number of ways, and is a key concept in evolutionary biology and quantitative genetics.
What are the typical methods (experimental designs) used to estimate this variance? How are they done?
I found papers such as this, which use paternal half sib breeding designs and the fit a model to estimate genetic variance and then multiply the values by 4 to estimate the G-matrix.

Of particular importance is the sire-level covariance matrix because, when multiplied by four, it provides an estimate of G.

I am interested in this because I am working on hemiclones and want to think about the differences, strengths, and weaknesses of classical methods compared to hemiclonal analysis.
*note, I mean to estimate the standing additive genetic variation within a population

I think for that you have to first establish the additive nature of the set of genes (i.e their independence).

The question is very much related to how we can estimate heritability and an answer will probably talk about regression of parents on offspring, identical twins, common garden and response to selection. The response to selection experiment intuitively seems to me is the most accurate and straightforward solution in order to measure the additive genetic variance.


text:
Molds and fungi usually found on book pages: identifying and preventing further spread
I know I can use bleach to kill most of the spores that live on my walls. I know I have to keep humidity below a certain level and temperature above another to prevent the spread of mold. I have read tons of websites on how to treat 'wall infestations'.
It's the mold that affects my books that worries me the most.
Especially now that I've discovered a very enticing market for used books. You can find bargains but sometimes the books smell "of mold".
I would like to know more about the kind of molds (and/or fungi or other bio-monsters) that usually feast on paper. The paper that makes the pages of a book.
Can someone point me to any kind of resource with answers to the following questions?

What are the most common biological causes of page deterioration?
1b. Can they be identified without lab instrumentation?
Can they spread from one book to another, and if yes how and how fast?
Can they be neutralized in some way (stopping the spread), or should one just burn the moldy books to avoid contagion?

I seek knowledge to protect knowledge.

Mold can grow in Extreme environments: they can live in very low water activity level, sometimes as lower as 0.71, live in refrigerator temperatures, these characteristics make it difficult to kill them thoroughly.
You can search "paper deterioration" in Google or library, then you can find lots article about this.
1) Causes
Foxing and other white, fluffy coating or light yellow powder on paper caused by: Eurotium rubrum, E. repens, Aspergillus versicolor, Penicillium solitum, and Penicillium decumbens. (Karbowska-Berent, Joanna, Joanna Jarmiłko, and Jolanta Czuczko.
Source: "Fungi in Fox Spots of a Drawing by Leon Wyczółkowski." Restaurator. International Journal for the Preservation of Library and Archival Material 35.2 (2014): 159-179.)
Black spots: Monilia, Aspergillus, Penicillium, Mucor, Stemphylium, Hormodendrum, Fusarium, Chaetomium, Byssochlamys
Source: Beckwith, Theodore Day, Wilbur Harold Swanson, and Thomas Marion Iiams. "Deterioration of paper: the cause and effect of foxing." Publ. Univ. Calif., biol Sci. 1.13 (1940): 299-356.
2) Spreading
For their spread, I think the mold can spread everywhere, but i did not find any resource about it.
3) Method that kill the mold
Drying the affected paper objects and applying 70% ethanol are the most preferred options to stop active fungal growth.
Source: Sequeira, Sílvia O., Eurico J. Cabrita, and Maria F. Macedo. "Fungal Biodeterioration of Paper: How are Paper and Book Conservators Dealing with it? An International Survey." Restaurator. International Journal for the Preservation of Library and Archival Material 35.2 (2014): 181-199.

source: Stack Exchange

license: Creative Commons - Attribution Share-Alike - https://creativecommons.org/licenses/by-sa/3.0/

sort: votes


text:
Clonidine's adrenonergic nature?
I am little confused here.
I used the term adrenoagonist and sympatholytic to describe the compound.
However, my teacher says that the correct term here is adrenomimetic -term.
My understanding of Terms

Adrenoagonist says that to bind to adrenoreceptor which prolongs the effect of NE in receptor.
Sympatholytic - inhibition of postganglionic fibers of sympathetic nervous system
Adrenomimetic - mimic effects of sympathetic nerve stimulation on pre- / postganglionic fibers.

I still think clonidine is sympatholytic i.e. adrenolytic (not adrenomimetic).
How can clonidine be adrenomimetic?
Why not adrenolytic?

Clonidine is an agonist on the α2 receptor... but then again norepinephrine is also an agonist on the α2 receptor. Then the physiologic ligand of the a2 receptor, norepinephrine is autocrine (meaning it is released from a cell then binds on a receptor on the same cell) causing negative feed back and it inhibits further release of norepinephrine from the neuron thus preventing overstimulation. Clonidine is quite specific to the α2 receptor and it doesn't really stimulate α1/β1/β2. Given a situation  where only the α2 receptor is stimulated and none of the other epinephrine receptors, you would have a net decrease of sympathetic effects since the amount of norepinephrine being released is begin decreased by the stimulation of the α2 receptor on the presynaptic membrane of the sympathetic neuron. Hence clonidine is an α2 agonist that is sympatholytic
I have no clue what the difference are between these

agonist vs mimetic
adreno vs sympatho

So I propose that clonidine is not adrenomimetic but adrenolytic (i.e. adrenonergic and sympatholytic in one word). Do you agree? - Agonist mean to bind etc adrenoagonist to bind to adrenoreceptor. Mimetic means positive ANS effect and/or mimicing the behaviour of the stimuli from ANS. Adreno - just adrenoreceptor where the neurotransmitter is acetylcholine. Sympatho is about if the drug has sympathetic effect and its nature (sympathopositive (-mimetic) vs sympathonegative (sympatholytic i.e. decreasing concentration of acetylcholine etc in case of adrenoreceptor in a synapse)).

I suppose I failed to mention this, but agonist means that it not only binds (as an antagonist binds as well,  both agonist/antagonist bind to either the receptors active site or an allosteric Site, depending on if it's competative/noncompetative) but it stimulates the receptor in a way that mimics the endogenous ligands response. Now the a2 receptors response when stimulated is adrenolytic.

Yes, you are right! Thank you for your answer!

I want to nuance the analysis presented above. Clonidine acts both directly on the alpha-2 receptors of blood vessels (triggering vasoconstriction), and then on the neurons alpha-2 receptors when it gets through the blood-brain barrier (inhibiting norepinephrine release). I think those definitions are too simplistic to fully qualify clonidine action. First reaction is sympathomimetic, prolonged action is sympatholytic.

Wikipedia seems to confirm what you have said. The a2 receptors are found not only in the presynaptic membrane of adrenergic neurons (and in the CNS), but also the vascular smooth muscle. Then there are the 2nd messengers involved, a1 acts via Gq (increasing IP3 and DAG) and a2 acts via Gi (decreasing cAMP). As I understand it, cAMP levels have varied effects on vascular smooth muscle (vasoconstriction/vasodilation) depending about which place int he body we are talking about, but if a1/a2 both worked on cAMP but in contradictory capacities then I would say no, but one is cAMP one is IP3/DAG.

all_licenses: Creative Commons - Attribution Share-Alike - https://creativecommons.org/licenses/by-sa/3.0/


text:
To understand side-effects for nebivolol's beta1 selective pathway in PubChem?
Assume you have a drug nebivolol.
When nebivolol is used as beta1 selective drug, beta2 is mostly for side effects, but this is not clear from PubChem.
I do not know any cases where nebivolol is used as beta1 and beta2 selective in the treatment.
Its first two pathways of interaction from PubChem.
Assume this pathway to be selective for some indication

and this nonselective (where side-effects stems mostly with large doses)

There no such differences in this section.
No explanations when side-effects come in each case of treatment plan (dose).
How can you understand side-effects of this adrenoblocker from selective beta1 pathway in PubChem?
Like in which cases beta2 selection becomes possible when beta1 is selective.
When beta2 is nonselective when beta1 is kept selective?
In other words, how can you read pharmacodynamics of this adrenoblocker in PubChem?
I.e. what the drug does to the body at any given dose.
I think there is little pieces of informations about it.
Implicitly, the side-effect can be read from Mechanism of Action:

where it refers at high doses to bronchospasm.
However, I would refer explicit discussion about side-effects and what we are trying to find and what are suspected as side-effects.

I don't understand the question. If you're talking about clinical effects, the distinction between "effects" and "side effects" depends on patient characteristics and the goal of therapy, so it would not be possible to differentiate these in a chemical database. If you're talking about something intrinsic to the molecule....I don't understand the question.

@Susan Thank you for your comment! Nebivolol is beta1 selective. Beta2 is only for side effects, but this is not clear from PubChem.

Again, if you're talking about clinical effects, I beg to differ that any particular effect or receptor activity can be labeled exclusively an effect or a side effect. For instance, the beta2 antagonism is likely responsible for the efficacy of these drugs to treat essential tremor, so  your statement beta2 is only for side effects doesn't seem correct.

The information that you have shown as a snapshot of pubchem is insufficient to comment about side effects.

How can you differentiate between effects and side-effects of this adrenoblocker in PubChem?

I think PubChem will list all sites (receptors) at which a drug reacts, rather than the preferred sites, which I think is appropriate.
The pharmacological properties are listed in Section 8: Pharmacology and Biochemistry:

Pharmacology
  Bisoprolol is a competitive, cardioselective β1-adrenergic antagonist. Activation of β1-receptors (located mainly in the heart) by epinephrine increases heart rate and the blood pressure causing the heart to consume more oxygen. β1-adrenergic blocking agents such as bisopolol lower the heart rate and blood pressure and may be used to reduce workload on the heart and hence oxygen demands. They are routinely prescribed in patients with ischemic heart disease. In addition, β1-selective blockers prevent the release of renin, a hormone produced by the kidneys causes constriction of blood vessels. Bisoprolol is lipophilic and exhibits no intrinsic sympathomimetic activity (ISA) or membrane-stabilizing activity...

From this, one cal tell that if there is activity at a beta-2 receptor, it is not the major intended site.
PubChem is a valuable resource, but it's not a one-stop-shopping site. That's the reason there are extensive references given for sites of interaction. It's a great starting place for investigation of a drug.

Thank you for your excellent answer with details! I completely agree with you that you need to consider the pathways of interaction parallel with Pharmacological properties to understand the selectiveness. It would be great if the selective pathways would be pinpointed explicitly, and those nonselective/selective corresponding to side-effects (often only with large doses) separately. My teacher says that use SmPCs to understand better PubChem. Do you know any source which collects SmPCs in one place?

@Masi - I'm afraid I don't know what SmPCs are. :(

SmPCs are Summaries of Product Characteristics found in most packages of drugs. Here is European guideline for them http://ec.europa.eu/health/files/eudralex/vol-2/c/smpc_guideline_rev2_en.pdf

US is enough, since I need to follow AAMI standards in my work that means US is the best choice. Thank you for the comment!  Is this the resource which you use http://www.pdr.net/ ?

@Masi - Yes, that's the one. Try to get your hands on a hard copy before subscribing, to make sure it will suit your needs.


text:
Suggestions for self-studying experimental techniques
I would appreciate any suggestion regarding self-studying experimental techniques used in biology for a theoretical scientist (candidate).
Maybe I should be more specific. I'm a biology major, but took as little experimental courses as possible, and I didn't paid much attention to those I took. (shame on me, right). Then I got a MS in applied math. When I revisit my molecular biology/genetics and biochemistry textbooks, I realized that they are mostly descriptive.
An experiment/technique oriented textbook would be very helpful for me, especially if it will also provide an overview of the methods.
My purpose is not the make up for the lost undergrad years, change my research orientation or master experimental skills. I'm aware that this is not achievable by reading some texts. My sole purpose is to not get lost on theory/experiment interface. When someone asks how something could be proven experimentally, I want to know what options are there in the field.

and I also wonder, why there is no self-study tag on biology.SE

Are you mostly interested in molecular biology oriented methods?

yes, especially genome sequencing and transcriptome quantifying methods

My undergrad mol. biol. book showed a lot of the original experimental data for establishing mol. biol. mechanisms, but it's pre-shotgun sequencing. It doesn't have a lot of the new, PCR-based stuff, because much of the basic biology was worked out before then. Should I hunt down that textbook title?

thank you, but I think I need something which covers also moder techniques.

I cannot more highly recommend Molecular Cloning 4 ed.. It is a really comprehensive text which covers numerous molecular biology experimental techniques including different types of PCR.
EDIT: Also, there are some great suggestions for other books and websites that also cover molecular biology techniques in the answers to this question: Short, concise, practical manual for doing experimental biology


text:
How to manage fat-gluten oxidation in celiac disease?
Assume you have been in non-gluten diet 20 years.
You built mass 8 mass with substances containing gluten (endogenous gluten stored in fat).
Then, celiac disease is diagnosed.
You have been 3 months in gluten free diet.
After mass period, you go to fat burning period.
The challenge starts: endogenous gluten stored in fat.
It is painful if too intensive training program.
Even little amount of gluten causes enteropathy.
What are the protectant methods for burning fat containing gluten?
How can you prove that there are no gluten-derived peptides that accumulate somewhere?
Since the cellular receptors for gluten have been identified, I think this may be over-complicating things. - Ryan's answer
I have not found any good solution for this, since I do not understand the pathophysiology enough well.
Toxic factors related to celiac disease

gliadin
...

How is endogenous gluten released from fat?

Do you have any evidence that gluten is stored in fat cells? As far as I know, it can only be synthesized by certain plants...

@MattDMo No. This was just a wild conjuncture. Probably, better one is gluten staying on the fat cells, but this would not apparently last long.

Gluten is a protein. The textbook answer is that protein is not stored in fat cells.
Proteins are hydrolyzed into amino acids through digestion; some amino acids are ketogenic, and can contribute carbon towards fat biosynthesis. So some of the carbon that originates in gluten may be incorporated into a fatty acid. But the carbon has no 'memory' of its previous role in a gluten molecule, and could not act as gluten when the fat is subsequently oxidized.
One can imagine an undigested gluten molecule (or its peptide fragments) binding to the exterior of a fat cell, but there's no mechanism for it to get incorporated into fat as an intact protein.

Thank you for your answer! I think you are correct. Is there any other way how gluten can remain in the body, like in some compartments? I think this is possible in stomach itself for some hours. I approximate that enough-digested-gluten-for-symptoms remain in the body at least 5 hours or better said its effect. Of course, this is relatively for each individual. To know where gluten stays would help to think better the mechanisms of gluten interaction with the body.

"Is there any other way how gluten can remain in the body, like in some compartments?" I think so, as I allude to in my last sentence. It's an interesting enough question to stand on its own. A problem is that the question is structured as an appeal to ignorance...how is one to conclusively prove that there are no gluten-derived  peptides that accumulate somewhre? Since the cellular receptors for gluten have been identified, I think this may be over-complicating things.

Thank you very much for your excellent comment! It is true. There must also be other aspects in this fat burning where gluten may remain stored.


text:
What does "chlorophyll photosynthesis peak" mean in relation to photon wavelength?
When reading about how green / leafy plants work,
I saw that they have chlorophyll A & B, which allow the plant to use the energy from light by capturing and transforming.
When reading about Chlorophyll, I saw some charts showing "peak points" for each type for light wavelengths. Apparently, each chlorophyll type has both peak points.
Since from further reading I learn that plants can use all the light in the visible spectrum for photosynthesis, I'm wondering (my questions >>):

What do "peak points" of wavelength correspond to / stand for?
How does light peaking in red spectrum (say around 630nm) differ to (or affect plants) compared to light in red spectrum peaking at 680nm (which I gather is Chlorophyll A peak-ish)?
When same amount of radiant flux is projected to leaves on different wavelengths, one far from a chlorophyll peak the other near, which one gives more energy to plant and why?

possible duplicate of What types of light can't a plant photosynthesize in?

I don't think it's a duplicate as the current question is quite specifically on the two peaks in the absorption spectra in chlorophyll.

First thing to note is that the absorption spectrum of chlorophyll A and chlorophyll B is similar but not the same. The reason for the appearance of absorption spectra is because the light is composed of photons of light.  Different photon has different  oscillation frequency and energy. The essence of Chlorophyll absorption is absorbing photons. After chlorophyll absorption of a photon, the electron transition from the ground state to the excited state, and get water electronic from the water cause decomposition. Only phonton that has resonant vibration frequency with electrons, and can meet the energy requirements of electronic transitions in order to be absorbed by chlorophyll. The Photon energy of different wavelengths of light is different. This is also the reason of different wavelengths of light affect chlorophyll light absorption.
Simply explain the absorption spectrum:

280 ~ 315nm  has small effect of plant morphology and physiological processes
315 ~ 400nnm absorption rate is small, impact the photoperiod, prevent stem elongation
400 ~ 520nm (blue) Absorption ratio of of chlorophyll and carotenoid in this light is maximum, it has the greatest impact on photosynthesis
520 ~ 610nm absorption rate is not high
610 ~ 720nm (red) low absorption rate.  It has a significant effect on photosynthesis and light cycle effect of  chlorophyll.
720 ~ 1000nm Low absorption rate, stimulate cell elongation, impact of flowering and seed germination
1000nm converted into heat

p.s. white light is a mixture of light

Hi @meateatingbunny, +1 on rephrasing and simplifying please. Also, can you please make 3 points to answer three parts of my question? Would greatly appreciate it! Thanks.

@meateatingbunny - why do you say that 610 ~ 720nm absorption rate low when all charts show differently? What do you mean, can you please explain?

Lets start with the absorption spectrum first (image from the Wikipedia page in chlorophyll):

What you see in the figure is the absorption of light thoughout the visible spectrum by chlorophyll a (blue) and b (red). The higher the peaks get, the higher the absorption is. What we can see, is that chlorophyll absorbs light roughly until 500 nanometers (nm) and from 620 nm. The light between this is not absorbed, but reflected. This is the green light and the reason why leafs are green. Above 700 nm the light is reflecxcted again, this is the reason why green leafs are visible white in infrared photography.
The peaking of the both chlorophylls ensures that light of the wavelength can be used efficiently for photosynthesis. Since leafs contain both chlorophylls, this gives a wider range for energy absorption than with only one. Additionally both absorption maxima are far enough from each other to prevent one chlorophyll from "stealing" light from the other.
The light closer located to the peak transmits more energy because it will absorbed better. The closer you get to the maxima of the absorption curves, the higher the absorption and the energy transfer will be.

What do "peak points" of wavelength correspond to / stand for?

The peak points of chlorophyll and their correspondence to the action spectrum indicate that the Chlorophylls play a major role in photosynthesis, it is also clear that chlorophyll isn't the only chemical involved in light harvesting and that other antenna pigments are utilized.
As mentioned in Chris' answer chlorophylls (plural) absorb and reflect light to a various degree at different frequencies, as explained on webpage in the link that was provided (in the section photosynthesis).
From Wikipedia's Chlorophyll webpage, "Measurement of chlorophyll content" section:

"In 90% acetone-water, the peak absorption wavelengths of chlorophyll a are 430 nm and 664 nm; peaks for chlorophyll b are 460 nm and 647 nm; chlorophyll c has a few isomers, peaks for c$_1$ are 442 nm and 630 nm; peaks for chlorophyll c$_2$ are 444 nm and 630 nm; variety c$_3$ is recently discovered; peaks for chlorophyll d are 401 nm, 455 nm and 696 nm;" chlorophyll f (also a newer discovery than information offered on Wikipedia) absorbs at 720nm - making it the most red-shifted chlorophyll to date.

Notice that chlorophyll A's Soret peak provides the greatest rate of growth. Blue light also affects phototropism more than red. Different wavelengths affect the plant differently, some cause growth while other wavelengths regulate bending, even seed development.
Here's an image from cell.us's webpage: "Action Spectrum Green Plants":

Note that plants reflect infrared light beyond 700nm, that is why they appear white (instead of black) in the right side infrared photograph:

Click to zoom

How does light peaking in red spectrum (say around 630nm) differ to (or affect plants) compared to light in red spectrum peaking at 680nm (which I gather is Chlorophyll A peak-ish)?

Chlorophyll b simply expands the range of usable light available to the plant.

When same amount of radiant flux is projected to leaves on different wavelengths, one far from a chlorophyll peak the other near, which one gives more energy to plant and why?

Light frequencies closest to one of the peaks has the most affect on growth, as you can see from the action spectrum blue light is more energetic than red and even mid-band (green) light has a small but significant effect on growth. Because green light is not absorbed some is reflected and some passes through the upper leaves to the lower shaded leaves.
Plants use light inefficiently because it isn't a limiting factor in its growth, instead CO$_2$ is.

all_licenses: Creative Commons - Attribution Share-Alike - https://creativecommons.org/licenses/by-sa/4.0/


text:
What is the charge on oligonucleotide 5' pGpGpApCpT 3' at pH 7.00?
What is the charge on the nucleotide 5'pGpGpApCpT 3' @ pH 7.00?
I thought adenine has 1 NH group and Guanine has NH and OH groups and cytosine and thymine  has OH groups on it and phosphate at 5' end an OH at 3' end .so at pH 2.00-3.00 OH will give away an electron and becomes negatively charged.so is it +2+2-1-1-1 which is -1? please explain?

Does gp mean group? Also, atoms don't become negatively charged by giving away electrons.

@canadianer GGACT are the nucleotides and the p stands for the phosphate backbone I presume..

@TheLastWord No I was referring to something  else that was present before the post was edited.

@canadianer k..

I didn't quite understand where you're getting all your OH groups, maybe from the enol tuatomers of the nucleotides. I tried to look up the pKa's for the keto tautomers of the nucleotides on Wikipedia1,2,3,4.

In general, if pH is below a functional group's pKa, that group will be protonated, and if pH is above pKa, it will be deprotonated, though this is an equilibrium.
If we assume pH of 7 and assume I have the pKas right ( these values are for the free nucleic acids, no deoxyribose attached ), then the primary amines on A, G, and C will be protonated NH3 groups with positive charges.The amides on G and T should remain deprotonated and uncharged.
So with the sequence GGACT, we have 4 primary amines, so 4 positive charges. However, we also have 5 phosphates, each carrying a negative charge, so +4 plus -5 is -1.
Of course all of this goes out the window once we bind that oligonucleotide to another oligonucleotide, I assume the hydrogen bonding will alter the pKas and charges. For all I know, simply attaching the nucleic acids to a deoxyribose and stacking them together changes the pKas.

we also have -ve charge at 3' oh end right?

I don't think the OH group on the 3' terminal deoxyribose would deprotonate, hydroxyl groups don't usually deprotonate at neutral pH unless something is pulling electrons away to stabilize the negative charge, such as the carbonyl in carboxylic acids. For example phenol has pKa of 9.99, while benzoic acid has pKa of 4.2.

There is no positive charge on the bases at pH 7, but there are 4 phosphodiester bonds which have -1 charge on each phosphate so total -4 also the terminal Phosphate at 5' end is free so it has a charge of -2.
Thus the overall charge of the oligonucleotide is -4-2=-6 (determined by number of phosphate groups)

There is no positive charge on the bases at pH 7, thus the overall charge of the oligonucleotide is -4 (determined by number of phosphate groups)... See details here: https://books.google.cz/books?id=v9HL5VyRmZcC&lpg=PA204&ots=DQYKZ9PxXg&dq=why%20amino%20group%20of%20nucleo%20bases%20protonated&pg=PA204#v=onepage&q=why%20amino%20group%20of%20nucleo%20bases%20protonated&f=false

It's unclear which text you are referencing in the linked book.  Can you quote the relevant portion?

Welcome to Biology.SE! Answers are much more likely to receive a favorable response if you include supporting references (primary literature is best) that other users can access — note the language this site uses is English. Without that support, your answer is indistinguishable from opinion. A good example of how to format references. ——— You may also want to take the [tour] and then consult the help pages for additional advice on [Answer] effectively on this site. Thank you!


text:
Flicker fusion threshold for insect eyes
I searched on bio-numbers for flicker-fusion threshold of insect eyes in general and for flies in particular. Ruck, 1961 gives values for some flies. Is there any database maintained for these values? Is there any comparative study done for many flies/insects?

"The flicker fusion frequencies of six laboratory insects, and the response of the compound eye to mains fluorescent ‘ripple’" (Miall, 1978) touches on flicker fusion thresholds for six different insect species commonly used in laboratory settings:

Locusta migratoria, Periplaneta americana, Saturnia pavonia, Antheraea pernyi, Glossina morsitans and Drosophila hydei.

I also found this paper: Potential Biological and Ecological Effects of Flickering Artificial Light (Boyles, 2014) that includes an exhaustive literature search of flicker-fusion thresholds and other related data for many species (including insects). If you're looking for an authoritative source, I'd say that this is it (as there doesn't seem to be a database with this information).


text:
Are there any mammals whose males can feed with milk?
Are there any mammals whose males can feed the offsprings with milk from the breasts?

@GriffinEvo: Birds are not mammals, meaning the question may still be valid.

See the answer to the question, it covers mammals too. @ChrisStronks

I edited the title of the linked Q so that it is clear that it also covers mammals.


text:
Why only female mammals have well developed mammary glands?
In course of evolution (possibly), mammary glands became vestigial in male mammals, but became fully developed in females. Is there any plausible explanation for this characteristic?

what exactly do you mean by plausible explanation?

Male mammary glands aren't vestigial, they're embryologically related to female mammary glands. So the question is "why are mammals sexually dimorphic" which gets tautological because the clade is defined in terms of mammary glands in the first place.

@Ryan Don't provide answer in comments. You can post it as another answer (saying "addition") if you provide some extra points that are not mentioned in previous answers. Use comments only for requesting clarification or providing suggestion related to post (not content).

Perhaps due to the close hormonal interplay between pregnancy and lactation it is more efficient to have the females generate milk. To let a male start to lactate a new mechanism needs to be developed in evolution, because a continuous 'just-in-case' lactation is energetically cumbersome. Developing new characteristics is not favored in evolution. Rather, coupling of a lactating mechanism to pregnancy (breast tissue growth) and labor (start of lactation) seems an evolutionary more parsimonious way of regulating lactation.

Just a note - they're not fully vestigial. Male lactation occurs in some species, including humans (the same mechanism that starts lactation in females still works for males). The Dayak fruit bat in particular has full blown lactating males. But yeah, taking mammals as a whole, these are relatively tiny exceptions. If you care about humans, though, yup, males can lactate just fine (though I have no idea about the amount or quality of milk so produced).

It comes down to investment in offspring. Males invest little in offspring only a little sperm whilst females have to develop the more expensive egg and the energy used through its development. In the mammalian line, this investment has increased in amount, initially through viviparity and then through the secretion of feeding fluids.
In species, such as humans, where there is a lasting pair bond and both male and female invest in rearing the young then I suppose there could be a situation where it makes equal sense for both males and females to nurse - although the cost of mammaries on other functions may favour specialisation anyway - but remember that, in many mammals, it is the mother alone that rears the offspring.
You may also find the answer to this question useful: Do any birds beside the family Columbidae (or any reptile or mammal) feed their young “Crop Milk”. What I hadn't realised is that there actually two species of bats where the male lactates.

It is not generally so that males invest little in offspring, especially not in humans. In fact, it is hypothesized that the rapid growth in brain volume in homo was made possible, at least in part, through the contribution of males in rearing the offspring (by gathering food etc.). Moreover, the 'more expensive egg' can be debated, as the vast amount of sperm cells generated during a male's lifetime vastly outnumbers the resources needed to ripen  the relatively small amount of eggs during a female's life.

I addressed your first point in the second paragraph. Male investment is high in humans, it is not in mammals in general and the dimorphism in mammaries did not evolve in humans. As to your second point, it's certainly true that males invest in reproduction to similar levels to females but in the individual offspring the investment is higher in the female.

The mammary glands in human males is not vestigial but rudimentary.
Human females have XX chromosomes and males have XY chromosome,here the "Y" chromosome is provided by father and "X" by the mother. At the time of fertilization, the father can provide either X or Y chromosome. If he provides Y chromosome the child will be male and if he provides X then child will be female. The mother always provides X chromosome. So the Y chromosome in the male child is responsible for the development of male sex organs and due to the presence of X chromosome the female sex organs are suppressed. Note that the female organs are suppressed but they are not eliminated. At the time of puberty males don't receive hormones which initiate the development of female sex organs. As a result the mammary glands stay in males as nipples but the hormone effects are only present in females and cause the formation of large amounts of fatty tissue near the nipples and develop as breasts.

A distinguishing characteristic of the class Mammalia is the presence of mammary glands. The mammary glands are modified sweat glands that produce milk, which is used to feed the young for some time after birth. Only mammals produce milk. Mammary glands are most obvious in humans, as the female human body stores large amounts of fatty tissue near the nipples, resulting in prominent breasts.
  http://en.wikipedia.org/wiki/Female

.

While estrogens are present in both men and women, they are usually present at significantly higher levels in women of reproductive age. They promote the development of female secondary sexual characteristics, such as breasts.
  http://en.wikipedia.org/wiki/Estrogen

The last sentence in your wikipedia quote is contradicting the first sentence in your answer.

@fileunderwater: I saw it but it may be because they just used "vestigial" as an inappropriate organ(literal sense)they may not be thinking the controversy in using "vestigial" and "rudimentary".

I rather see it as a reason to have healthy scepticism towards unreferenced statements in Wikipedia. But the reason is probably, as you say, how they define 'vestigial' (I don't think human nipples should be labelled 'vestigial' using normal definitions of the word though). It makes for a contradiction in your answer though.

@fileunderwater:I just edited the answer and removed the portion, but I think its unfair to remove that portion, but it is for a good reason.Also you are thinking that it shouldn't be labelled as "vestigial". I have learned many things from your past comments and I am hoping your assistance will improve my knowledge and my approach towards SE.Thank you !!

I just added [citation needed] to the WP article's sentence in question.

I think this answer, while of decent quality, does not actually address the question the OP is asking. This is the mechanics of how there is a difference not the reasons why there is a difference.

Incidentally, the XY system is not used in all mammals so there is a slight lack of generality here.

@JackAidley:From my narrow knowledge in the field I tried to give an answer, do science know the reason why there is a difference ?


text:
human anti-mouse antibody
I have heard about human anti-mouse antibodies (HAMAs) and read that HAMAs neutralize murine antibodies, therefore decreasing the effectiveness of those murine antibodies. Is this true that HAMAs neutralize the murine antibodies? How?

@user137 please don't answer questions in comments. Write a full answer instead.

@MattDMo It's fixed, just took me a while to get the protein sequences aligned. I don't like to make answers without some evidence.

@MattDMo - I'm curious, this is second comment regarding answering in comments I've seen today. I wasn't aware this is normal policy. Personally I often leave a brief answer in a comment if I don't have time (or patients) to write a full good quality answer - I just put the comment in, if someone wants to take it for an answer thats fine by me. Perhaps there needs to be a meta post to discuss policy? (I don't know the extent of the answer that was left as a comment here, but certainly on the other I saw a user was told to put a quick one line comment in to a proper answer)

@GriffinEvo see this meta post for a previous discussion on the phenomenon, and of course this recent related post as well. Regarding the (now-deleted) answer-as-comment here, it was probably 6 or 7 lines long, and covered all the points in the answer below, with the exception of the alignment.

As mentioned in the comment, all antibodies have constant regions and variable regions. The variable regions are the binding sites on the ends of the two arms, and the constant regions are the rest of the molecule. When humans are injected with murine antibodies, the immune system recognises and sets up an immune response to them, through generation of HAMA. This affects the efficacy of the antibody because it is attacked the same way a pathogenic substance would be. Antibodies have different effects, but the most important one in this instance is through opsonisation (http://en.wikipedia.org/wiki/Opsonin#Examples). Because antibodies have two arms, they can bind multiple targets and cause a grouping and clumping. This also causes activation and phagocytosis through phagocytes (generally macrophages) which will ingest and destroy the therapeutic antibodies.
Antibodies that are mostly human with just mouse variable regions have been created to try and counter this problem, known as chimeric antibodies. Humanised antibodies are also available, which are entirely human with only the very tips of the variable regions (known as the hypervariable regions) are from mice. This reduces the problem as far as possible (http://en.wikipedia.org/wiki/Fusion_protein#Chimeric_protein_drugs).

It's common for the human immune system to create antibodies against many proteins, even some human proteins. Hemophiliacs who receive regular doses of clotting factor proteins often develop neutralizing antibodies against the clotting factor proteins, even though they are a human protein1.
Therefore it's not surprising that antibodies would be developed against mouse antibodies. There are differences between mouse and human antibodies, and human antibodies may bind to these different regions. I found the amino acid sequences for human2 and mouse3 immunoglobulin gamma heavy chain constant region and aligned them by hand and marked the matching amino acids:
Human: MELGLSWVFLVAILEGVQCEVQLVESGGGLVQPGGSLRLSCAASGFTFSSYWMSWVRQAPGKGLEWVANIKQDGSEKYYVDSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARECDNWF DPWGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKKVEPKSCDKTHT CPPC      PAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK
       |   |  |||| || |||||||||||||||| ||||  ||||||||||| | | |||||| ||||||| |       || | |||||||||||||| |  || ||| |||| ||||||      | |||||  ||||| |  ||| ||||    | |    |||||| |||| ||| |||| | | |||||| ||| |||  || ||||||    ||  | | |  | | |||| ||     |   ||||      ||| | |||||| |||  || |||| || |||||||||  || |   | |  |||| | |    | |||| |||| |   ||||  ||| |||| || || ||| |||| ||  | |||| |||    |    ||||||| || | || ||| ||   | ||| | |||||||| | ||||    | |     ||| | || | | |  |  | ||||
Mouse: MDSRLNLVFLVLILKGVQCEVQLVESGGGLVKPGGSRKLSCAASGFTFSDYGMHWVRQAPEKGLEWVAYINSGSTTIYYADTVKGRFTISRDNAKNTLFLQMTSLRSEDTAMYYCARELWLRRIDYWGQGTTITVSSAKTTPPSVYPLAPGCGDTTGSSVTLGCLVKGYFPESVTVTWNSGSLSSSVHTFPALLQS GLYTMSSSVTVPSSTWPSQTVTCSVAHPASSTTVDKKLEPSGPISTINPCPPCKECHKCPAPNLEGGPSVFIFPPNIKDVLMISLTPKVTCVVVDVSEDDPDVQISWFVNNVEVHTAQTQTHREDYNSTIRVVSALPIQHQDWMSGKEFKCKVNNKDLPSPIERTISKIKGLVRAPQVYILPPPAEQLSRKDVSLTCLVVGFNPGDISVEWTSNGHTEENYKDTAPVLDSDGSYFIYSKLDIKTSKWEKTDSFSCNVRHEGLKNYYLKKTISRSPGK

As you can see, there are a lot of matching residues, but several differences. Keep in mind that antibodies also contain light chains, and that there are several other types of antibodies, I just used the IgG heavy chain as an example.

is there a way to prevent this? Also, how does this neutralize the antibodies?

This could be prevented by using human antibody genes in cell culture instead of growing antibodies in mice and extracting them from the blood. As far as neutralization goes, the human antibody may just mark it for uptake by white blood cells.

user137's comment about opsonized particles being marked for phagocytosis by macrophages is right on. There are a number of ways that antibodies can be neutralized (e.g. steric hinderance), but being eaten up definitely does the trick.  However, growing antibodies in culture won't solve the problem. They are recognized, in large part, just by their Amino Acid sequence, rather than any culture conditions (you might make an argument for glycosylation). The variability of antibodies will always make them vulnerable to immune responses. There are even HAHA (human anti-human antibody responses.


text:
Is our color vision calibrated to sky, vegetation, and blood?
Our color vision is based on three types of receptors (cones) which are sensitive to three distinct locations on the spectrum:  420–440 nm, 534–555 nm, and 564–580 nm.  We label them "red", "green", and "blue", but these names are arbitrary.  However, their locations on the spectrum may not be arbitrary.
I notice that they correspond to the colors of the three most important things in our natural habitat:  sky (the above), vegetation (the below), and blood (the sign of potential food or alarm, depending upon whether you are the hunter or the prey).
Am I on to something here, or am I just dreaming in technicolor?

It is proposed by various groups that color vision developed due to its evolutionary benefits in enhancing the recognition of (ripe) fruits. I think it's all highly speculative but I can dig up some articles if needed. The evolutionary benefits of recognizing sky seems remote, especially for terrestrials. Recognizing the color of blood makes no sense as prey is most often not yet bleeding and wounds are recognized through pain. Rather, the 3 cone types and with color opponency enable the visual system to see many colors in the visible spectrum, that reportedly help to recognize fruit.

Fruit!  I hadn't thought of that.  I guess that does make a lot more sense than sky.  Thanks, Chris.

And most mammals don't have the receptors to see blue light. But flowering plants are relatively recent, do non-flowering plants make colored fruits?

Also, I think dogs and some other predator animals lack red cone cells in their eye due to never having the pressure to need to easily detect berries (often red). [Citation needed] I have never heard anything about needing to see blood as a sign of alarm/food. Emphasising what @ChrisStronks said, if an animal is bleeding so heavily that it can be seen at distance, odds are as a predator that your job is done. As for tracking/alarm, smell would and does serve much better for animals +doesn't require staring at the ground as much.

Short answer
Color vision is not based on a calibration to the sky, vegetation and blood. The current leading theory of the development of trichromatic vision in humans is based on the foraging of fruit in our primate ancestors.
Background
The places of red, green and blue wavelengths in the spectrum are physically defined and, therefore, not arbitrary; see this question on Biology SE.
Before continuing, it is good to mention that trichromacy (the presence of red, green and blue cones in the retina) is typical for primates. For example, dichromacy (<500 and >500 nm cones) is the most common configuration in mammals. The split in red and green photopigment in primates developed relatively late in evolution (30-40 mln years ago), inferring trichromacy in primates from a dichromatic ancestor. Note that many avian and aquatic species may be tetrachromats or up (4 or more photopigments) (Nathans et al. 1999), with the mantis shrimp featuring no less than 10(!) different photopigments (Cronin & Marshall, 1989).
It is believed that the red-green-blue system in primates has evolved due to its benefits for identifying fruit on a background of foliage, and to assess the ripeness of fruits. The food of monkeys typically consists of fruit, and young leaves. Young leaves are light-green, while ripe fruits are often yellow (e.g., bananas) and orange (e.g. oranges, mangos) (Osorio & Vorobyev, 1996). Unripe fruits typically contain chlorophyll and resemble the color of foliage (green). Hence, to discern ripe, nutritious fruits (yellow-orange or 570-620 nm) from foliage (green or 495-570 nm - frequencies mentioned were copied from wikipedia), it makes sense to have good resolution in the red-green area. The red opsin has its maximum at 530, and the green at 560 nm. The blue has its peak a lot lower at 425 nm (Nathans et al. 1999). Hence, the red and green opsins, which evolved late in evolution, are both right in the "ripe fruit area" and have absorption spectra very close together. Hence, there is a lot of color sensitivity (high color resolution) in the red-green area, which covers reds, oranges, yellows and greenish colors. This gives primates a very high sensitivity to recognize fruits, and especially ripe fruits in a green background. The resolution in the green-blue range is smaller (blues, purples).
As a background: color vision in trichromats works by color opponency, and specifically a yellow-blue and red-green color opponency. This means there exists no yellowish blue hue, or a reddish green (De Valois & De Valois, 1993). The red-green opponency means that we have exquisite sensitivity in the red-green pathway, as red cones suppress green cone output and vice versa. In other words, there is a high color acuity in the red-green area that helps to recognize fruits.
As a side note: The trichromat color vision scenario in primates supports the notion that cats (for example), being predators with dichromatic vision, do not benefit from being able to identify ripe and unripe fruits, as mentioned by @GoodGravy. In contrast, cats need excellent night vision and motion sensitivity, being nocturnal hunters. Indeed, color vision versus night vision, and color vision versus high motion sensitivity are both trade-offs that do not go hand in hand (Kelber et al, 2002).
References
- Cronin & Marshall, Nature (1989); 137-40
- De Valois & De Valois, Vis Res (1993); 33(8): 1053-65
- Kelber et al., Biol Rev (2003); 78: 81–118
- Nathans et al., Neuron (1999); 24(2): 299–312

Thank you, @ChrisStronks.  This is a wonderful answer.  I have heard about the mantis shrimp and its numerous color receptors.  Amazing!  It makes you wonder what their experience of the world is like.  I've seen multispectral and hyperspectral imagery.  It's an overwhelming amount of information.  Anyway, thanks again.  I'm now in the mood for some fruit...

Is our color vision calibrated to sky, vegetation, and blood? TL;DR; No
Normal person has 3 types of detectors in his eyes called cones that are responsible for color vision. Cones can be: red (only name), green, or blue. They do not respond to single color but rather to range of frequencies. Under few layers of specific cells they connect to optic nerve and nerve transmits signals to the brain. (This is simplified version as same nerves are responsible also for intensity) Based on intensity (how much nerve fires) of signals from different color nerves brain is capable of figuring out what color it really is. Every type of cone has it's responsive frequency range.

If you would look at the samples below of peak ranges, it makes it pretty visible that Red cones peak at what normal person perceives as yellow (it could be 👽 blood though :) ) and not red, while blue cones peak at very intense blue-violet that sky isn't, green is on the right track though.
So I must regretfully say that a very 'romantic' theory on vision calibrated to sky, vegetation, and blood cannot be right since 'red' cones in fact respond to yellow light.
Blue cones peaks

Green cones peaks

Red cones peaks

Here, you can find the tool that I have used to get RGB/Hex values from wavelengths.

Would downvoter care to elaborate where did I go amiss?


text:
How big should the human population be, as predicted by body mass?
I would guess there is a theory in biology which states that the population size for a given species is inversely proportional to the body mass of individuals in that species.  In other words, there are zillions of ants, billions of mice, millions of deer, and just a few elephants.
If humans existed in a "state of nature" (i.e., were not able to manipulate their environment more than any other mammal), what population size would be predicted by their ~70 kg body mass?
(Assume that the human population has spread globally.)
I imagine we would fall somewhere between the wolf and bear populations.

The question is interesting. Yes, there is probably such negative correlation. However, in order to make predictions from such regression, one has to make some arbitrary decision about what species to include in the regression. Do you want to include only mammals, all vertebrates or all living things (assuming this is well defined) on earth? I am hoping someone can come up with a good answer of what the regression look like depending on what clades we include in the regression.

Note also, that in general large species suffer from the presence of human and therefore, such regression is likely to be very different today to what it was a 1000 years ago.

It is certainly possible to use general relationships to predict human population density or abundance. The relationship between population density/abundance and body size is an old topic in ecology that falls within the field of allometrics (how different features of organisms scale with body size). A closely related allometric relationship is Kleiber's law, which deals with the relationship between body size and metabolic rate.
Your assumption of a generally negative relationship between body size and abundance is correct, and the relationship is approximately linear on a log-log scale. As mentioned in the comments, the exact shape of the regression will depend on what organism groups are included, since the scaling relationship is known to differ between taxa. However, to get a reasonable ballpark figure we can look at a the classic paper by Damuth (1981), which include 307 mammal herbivores. The scaling relationship found there is:
$$D = 10^{4.23}*W^{-0.75}$$
where density (D) is in individuals per km2 and body weight is adult body mass in grams. Using your example of 70kg for humans would result in a predicted density of 3.9 individuals/km2. The scaling coefficient -0.75 (or -3/4) is often labelled "Damuth's rule", and is usually also found when pooling both ectotherms and endotherms.
However, remember that this regression was based on primary consumers, i.e. mainly herbivore grazers. Alternative data for groups of mammals can be found in Peters & Raelson (1984). That paper presents regressions for both carnivores+omnivores as well as for all groups of mammals (herbivores, carnivores, omnivores). The relationship for carnivores + omnivores (CO) is:
$$D = 15.7*W^{-1.15}$$
and the regression with all groups is (HCO):
$$D = 40.7*W^{-0.859}$$
In both of these regressions body weight is expressed in kg. Using the CO regression results in a predicted human density of 0.1 individuals/km2, and the HCO regression gives a human density of 1.1 individuals/km2.
Looking at these values, it is reasonable to assume that the predicted "natural" population density of humans lies in the range 0.1 - 4 individuals/km2, but certainly biased towards the lower part of the range. I suspect that a regression looking only at primates would predict an even lower value, so I would not rule out values below 0.1 individuals/km2.
To get a grasp of what this means in actual population numbers, we can extrapolate the densities to the entire earth. If we use the predicted density from the carnivore+omnivore model in Peters & Raelson (1984) and the entire land area of the earth, which is ~150 million km2 (Wikipedia: Earth), the predicted human population would be about 18 million. Using the high estimate from Damuth (1981) would correspond to about 600 million people. However, remember that this is using the entire land area, including deserts, mountain ranges and arctic areas.
Another point of reference is the estimated historical human population density during the Paleolithic, when we lived as hunter-gatherers. I have often seen population densities in the area of 1 per mile2, i.e. 2.6 individuals per km2. These estimates are naturally very uncertain, but shows that the allometric scaling relationships seem to provide at least reasonable order-of-magnitude estimates of "natural" human population density.
It is also somewhat enlightening to consider that our current human population size of 7 billion corresponds to the predicted population size of a 380 gram carnivorous mammal (using the CO relationship from Peters & Raelson (1984)), making the unrealistic assumption that this mammal has managed to colonize all land areas on earth. This shows pretty clearly what an outlier modern humans represent.
The generality of these types of relationships for predicting densities of individual species can however be questioned. For instance, the regression relationships also differ between climatic regions (see Peters & Raelson (1984) for more), and, as indicated in the comment by @remi.b, it is also unclear what population states that should be included. For instance, should only population densities in "ideal" habitats be used, when we know that population densities of the same species can differ by orders of magnitude in different habitats? It is also unclear if/how to consider the effects of modern humans. Silva & Downing (1995) also argue that the scaling relationship is nonlinear on the log scale, with a more shallow slope for larger species, and also find essentially no relationship between body size and density within some groups of species. More recent papers have also highlighted that studies on global relationships between density and body size often mix-up several different processes and relationships, see White et al (2007) for an overview on this.

dang thats 2 billion people tops.

@shigeta but that is using the entire surface of the earth, right (including water)?

I think I googled the land area of the earth, but it would include antarctica.  I think it would be great if we could get to 2b human beings though.  with good technology we could be sustainable I think.

Thank you, @fileunderwater.  This is a very thorough answer.  I especially appreciate the equations!

@SlowMagic You're welcome. I enjoyed revisiting some of the litterature and writing it.


text:
How to inhibit formation of specific antibodies (to antisera)?
Is there a way to inhibit an antibody response to a specific antigen using immunosupression? I am interested in reducing the anti-antibody formation to animal antibodies such as murine antibodies in antisera.
I have read that there are ways to decrease the immune response to antisera but haven't found a detailed paper on how they work.
Update 12/3/2014: Would this work? http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2374127/pdf/0100491.pdf
Although it is streptavidin, could a similar process be used for antibodies?

Related post

Are you asking about in vitro or in vivo?

in vivo... if possible...

As far as I know there is no way, except for suppressing the immune response at all. If it were possible, this would be a great help against autoimmune diseases.

If you are referring to reducing the titer in the immunized animal, you could give the animal a drug that has immunosuppressive properties. Velcade (PS-341) is a drug that effectively reduces plasma cell counts. There are many immunosuppressive drugs available.
If you  want to reduce the antibodies affinity, you are undertaking a much more delicate process. Affinity can be transiently affected by introducing a  unfavorable environment for the antibody. You can accomplish this by changing the temp, salt conditions, pH or introducing a denaturing agent like guanidine HCL.
Changing any of these conditions in just a little excess could also result in a permanent reduction in affinity.
If you are attempting to tailor the affinity constant of the Ab by changing the architecture of the antigen, you are correct this can be done using site directed mutagenesis. However I think it would be more effective to perform a random mutagenesis and then screen the mutants for the Ka value you want.
Lastly the antigen could also be post-translationally modified. After performing in vitro transcription and translation you could modify your antigen using different strategies such as ubiquination, phosphorylation, prenylation etc... All these could potentially affect Ka as well.
A Hybridoma system to produce a monoclonal antibody should also be explored. Here is a reputable company that produces monoclonal antibodies:
http://www.sdix.com/Products/Custom-Antibody-Services/Ab-Production-and-Services/mAb-Antibody-Development/Hybridoma-Development.aspx

Hello @rhill45 , I did read about reducing immune response to an antiserum so I would like more info on that.. but otherwise, thanks for the info... BTW, can site directed mutagenesis and ubiquination, phosphorylation, prenylation etc. be used to change the Fc region of an antibody? Can it  also be used to chimerize the antibody?

@TAbraham check this out http://m.peds.oxfordjournals.org/content/23/4/289

Chimerize the ab?? To the antigen? What are you trying to do?

I am trying to decrease the immune response to foreign antibodies (this is why i was talking about anti-sera) so the antigen would be the foreign antibody...

Also, would creating antibodies against the antibody-producing cells work?

If you are creating an antibody against another antibody what you are creating is called a secondary antibody. If you want to decrease antibody production in a host the way to do that is through immunocompromising  the host. That are many ways to do this with varying specificity: radiation, chemicals, viral infections, giving the host certain cancers; even an autoimmune disease could work I guess. Let me know what I'm missing in my answer to get it marked. Using Cyclosporine is going to be your easiest and most well studied method.

I am trying decrease the production of secondary antibodies...

@T ABraham Let me know what my answer is missing to get marked as correct.

are there any other studied ways to decrease the production of secondary antibodies? It seems cyclosporin won't work... I want the most specific way to do so if possible... BTW, Thnx for your answer...

Velcade is a proteosome inhibitor that is very specific for lowering plasma cell concentration (these are the cells producing the antibody).  Have you looked at a Hybridoma system to make a polyclonal secondary antibody?

What do you mean about polyclonal secondary antibody? i know about hybridoma systems for monoclonal antibodies but don't fit my needs... I am trying to stay away from chimeric and humanized antibodies as well...

Oops meant monoclonal (sorry iPhone)

also, I don't want to use hybridomas because it would take away the reason for the project I am using this info for...

Let us continue this discussion in chat.

Would you like to talk in chat?


text:
How was birth control realized in ancient days?
In ancient days, there were no birth control methods like condoms or contraceptive pills available. How did they control pregnancy?

I have no reference to this at this time, but I believe the Romans used the intenstines of sheep as a condom.

Although the question is well viewed and upvoted, and has answers already I'm going to suggest that this is perhaps not the most suitable SE site - it's more history rather than biology: http://history.stackexchange.com/

The need for birth control has been a concept for thousands of years and remains an important issue for men and women today, the need for contraception helped humans to achieve control over our population and healthy sexual relationships without having the fear of pregnancy.
The medieval contraception methods mostly relies on plant based oral contraceptives and also plant based suppositories. There is always an element of myth in the usage of these medicines to control child birth, but due to the overusage of Silphion, the plant was gone extinct.

Silphion was a plant that grew in the hills near the Greek city-state of Cyrene located in North Africa. This plant was one example of an oral contraceptive in ancient times, based partly on Pliny's statement that it could be used "to promote the menstrual discharge" but since then has been harvested into extinction.
Reference

Asafoetida is widely used as a  contraceptive in medieval period, Asafoetida or asafetida is the dried latex exuded from the rhizome or tap root of several species of Ferula.

One of the earliest contraceptive pioneers was Soranus of Ephesus, a Greek physician of the 2nd century C.E. who practiced in Rome and wrote extensively on obstetrics, gynecology and pediatrics. Many of his texts exist to this day which describes plants that were recommended for use as oral contraceptives, suppositories and abortifacients: galbanum, silphium, opopanax, myrtle, rue, white pepper, cow parsnip and pomegranate.
Reference

Also condom is not a modern mechanism it was used in ancient times, but the usage was to protect oneself from sexually transmitted diseases, but an unknown chemical used  which is a spermicide and this resulted in contraception.

Records of condom use dates back to 3000 B.C. where King Minos of Crete, son of Zeus and Europa, utilized the bladders of goats to protect himself during intercourse.
In the 1500s, a syphilis epidemic spread across Europe. It was at this time that Gabriel Fallopius,created a linen
condom as a means to protect from the continuing spread of disease . This proved especially effective when soaked in an unknown chemical solution acting as a spermicide.
Reference

The medieval contraception methods includes,

Condoms
Female Barrier Methods
Herbs and Rituals
Intrauterine devices
Male methods
The Pill

These methods are widely described in this article.

Condoms have been around for a long long time. Supposedly, condoms made from such materials as fish bladders, linen sheaths, and animal intestines where in use around 3000 BC (reference). Coitus interruptus was tried before that which may or may not be successful. Infanticide was also practiced where an unwanted child was born in ancient societies (reference). You can read all about the history of birth control on wiki.


text:
How does anger relate to blood pressure?
Anger is an emotion generated by neural processes in the brain and is associated with elevated blood pressure. How can an emotion, which is totally related to brain, result in blood pressure changes?

Anger is a common emotion in most animals and it is highly related to stress. At time of anger body usually releases stress hormones and  the body's way to respond to stress is by sympathetic nervous system activation which results in the fight-or-flight response.

Anger is an emotional response related to one's psychological interpretation of having been threatened. Often it indicates when one's basic boundaries are violated. Sheila Videbeck describes anger as a normal emotion that involves a strong uncomfortable and emotional response to a perceived provocation.
Anger may have physical correlates such as increased heart rate, blood pressure, and levels of adrenaline and noradrenaline. Some view anger as an emotion which triggers part of the fight or flight brain response.
http://en.wikipedia.org/wiki/Anger

Stress and anger trigger the release of stress hormone Cortisol in the body. Mild releases of Cortisol can give the body a sudden burst of energy,it also may increase the heart rate to 180 times a minute and  blood pressure from 120 over 80 to 220 over 130.
Anger is processed in hypothalamus and amygdala,the roles of two peptide hormones, corticotropin-releasing hormone (CRH) is responsible for the production of Cortisol (Primary stress hormone) and  arginine-vassopressin (AVP) produces Vasopressin, which triggers blood pressure by increases reabsorption of water by the kidneys, this results in contraction of blood vessels which raises blood pressure.

The human stress response involves a complex signaling pathway among neurons and somatic cells. While our understanding of the chemical interactions underlying the stress response has increased vastly in recent years, much remains poorly understood. The roles of two peptide hormones, corticotropin-releasing hormone (CRH) and arginine-vassopressin (AVP), have been widely studied. Stimulated by an environmental stressor, neurons in the hypothalamus secrete CRH and AVP.
CRH, a short polypeptide, is transported to the anterior pituitary, where it stimulates the secretion of corticotropin (4). Consequently, corticotropin stimulates increased production of corticosteroids including cortisol, the primary actor directly impacting the stress response (5). Vasopressin, a small hormone molecule, increases reabsorption of water by the kidneys and induces vasoconstriction, the contraction of blood vessels, thereby raising blood pressure.
http://dujs.dartmouth.edu/fall-2010/the-physiology-of-stress-cortisol-and-the-hypothalamic-pituitary-adrenal-axis#.VH7k8cnQoX0


text:
What mechanism prevent Colchicum Autumnale from being affected by Colchicine
Colchicine is a chemical usually used to cause polyploidy mutant by inhibiting chromosome segregation. It is synthesized in the Colchicum Autumnale. But what mechanism prevent C. Autumnale from being affected by the Colchicine?


text:
allogeneic organ transplant and the immune response
What happens when an allogeneic organ transplant is performed on an immunocompetent recipient vs. when it is performed on an immunosuppresed recipient using a donated organ with immunocompetent cells?

Is this a homework question?

No, it's a question I saw when reviewing organ transplant

The situation is just as you'd imagine. In an immunocompetant recipient, a host-versus-graft reaction occurs. This is called transplant rejection. Single episodes (acute rejection) are easy to treat and rarely lead to organ failure. Chronic rejection is the leading cause of organ transplant failure. The organ slowly loses its function and symptoms start to appear. This type of rejection cannot be effectively treated with immunosuppressants. Some people may need another transplant.[1]
In bone marrow and other transplants, T cells in the allografts reconstitute T-cell immunity in the recipient. Unfortunately, these T cells recognize the recipient as 'non-self' and employ a wide range of immune mechanisms to attack recipient tissues in a process known as graft-versus-host disease (GVHD).[2]

Acute GVHD describes a distinctive syndrome of dermatitis, hepatitis, and enteritis developing within 100 days after allogeneic hematopoietic-cell transplantation (HCT). Chronic GVHD describes a more diverse syndrome developing after day 100. In addition to allogeneic HCT, procedures associated with high risk of GVHD include transplantation of solid organs containing lymphoid tissue and transfusion of unirradiated blood products.[3]

GVHD is increasing due to the increasing number of allogeneic hematopoietic cell transplantations (HCT). More than 25,000 allogeneic transplantations performed annually. Given current trends, the number of transplants from unrelated donors is expected to double within the next five years, significantly increasing the population of patients with GVHD.[4] Yet the major complication, GVHD, remains lethal and limits the use of this important therapy.
Long-term immunosuppressants are usually the treatment regimen for chronic GVHD. Fungal, bacterial, and viral infections are a major risk with this treatment since the immune system will be suppressed for a very long time.[5]
[1] Transplant rejection
[2] Graft-versus-host disease
[3] Graft Versus Host Disease
[4] Graft-versus-Host Disease
[5] Graft vs Host Disease: An Overview in Bone Marrow Transplant


text:
Theranos blood test's Specificity and Accuracy?
I am studying the limitations of Theranos device.
They have 238 documents in Google Patents Search (Inassignee as Theranos), here.
Which patents are relevant for their device?
They have 70 tests which are not necessary to consider now.
Just the basics of the device.
My intuition says that it uses PCR to multiply the microliter of blood for the analysis.
What are limiations of multiplying so little blood?
I think this is where their engineering work applies.
What components do you need to multiply?
I think all is not necessary.
Otherwise, the specificity and accuracy can be difficult to hold.
How can Theranos blood test's be specific and accurate compared to conventional full blood test?
I suspect if you can do the conventional full blood test with Theranos device with the same accuracy and specificity as the conventional one.

Which patents are relevant for their device?

They currently only have about 27 issued patents (US)
Of those patents about 10 of them are related to network/internet/health record/algorithm (so not really about their device)
The remaining patents focus on a few areas: sample acquisition (getting the blood drop and handling of it), Patch based technology (before they pivoted), sample quality analysis (this is key to making sure the small sample is good quality), microfluidics/modular device (these are the patents relating to the actual testing and device)

What are the limitations of multiplying so little blood?

From a molecular standpoint (PCR) the smaller sample you start with, the less of the original nucleic acid template you have to start with.  Lower end PCR testing uses 10uL of sample.  This is ok when you are trying to bring molecular diagnostics to the point of care or developing markets, but the best PCR tests use 50uL of sample (more than a drop of blood which is traditionally 25uL or so),  regardless of how good your device is, at some point you start amplifying junk. The better the primers and other reagents the less junk you amplify, but at the end of the day you want more sample to be more sensitive.
From a NON-molecular standpoint (chemistries, cellular analysis, coag, etc.), they aren't/can't multiply the analyte of interest.  In these tests, they rely more on dilution or small volumes to perform detection.  This is where their ability to reliably dilute and correct for the dilution comes into play. Although there is certain limits to this.  Coagulation tests would not be representative if you diluted the sample so in this case they will have to rely on working with a small volume.  Cellular analysis can only be diluted so much as you get to a point where you miss important but less common cell types.  Take eosinophils (a type of WBC) while they can get to 15 or 20 percent of all WBC cell types, in general they only make up a couple percent.  Assume that 2% of all your WBCs are eosinophils.  If you only have 3500 WBCs per 1uL of blood (which is on the lower end of normal), you have approximately only 70 of these cells in that same volume.  If you dilute the sample so that way there is more of it available for other tests like electrolytes, but you still analyze 1uL, The number of those cells is even less.  If you had perfect detection you might be ok, but you generally want to analyze more blood so that statistics work in your favor and your performance goes up. Most of the other chemistries, proteins, and as mentioned earlier, electrolytes will probably be ok for dilution and detection and in fact this is why there are plenty of small devices already out there that use a drop of blood and perform the same tests.
Their issue in diluting or amplifying analytes of interest is that biology still is governed by the laws of physics and even though technology may allow you to analyze samples down to 10nL, that just isn't use full in biology, and limits their their capability to perform the supposed menu of tests from one microtainer of blood.  They don't publicize that they may need to draw a venous sample (needle in your arm) for some tests and depending on what tests you order, you may need a separate finger prick for each test.

How can Theranos blood test's be specific and accurate compared to conventional full blood test?

If they reliably control dilution of the small sample, they maybe able to provide high quality results for some tests.
Finger pricks actually can be easily skewed via technique.  Theranos has specially trained people to draw these samples, but it is time consuming >5 mins and until it is fully automated this is one variable that no matter how good sensors are, you can't correct for variation pre-analytically when you have that small sample.  Finger pricks also skew certain tests even when technique is good such as platelet and WBC values in the CBC.
I discussed the sample volume issue above regarding PCR amplification and how good tests use up to 50uL.  So if they compare to a 10uL test, they may be able to compete, but not full lab quality.
A lot of the future techniques they have regarding small techniques (in their applications) like their coagulation tests work great in an academic setting, but they fail to be able to handle the full range of blood samples they will see, so it will be interesting to see how that plays out.

Takeaway

Theranos is working on modular micofluidic devices/cartridges that will use small sample
There are certain biological limitations that stand in the way of them performing all the tests in their catalog with only a drop of blood
The FDA will eventually be requiring them to submit their tests for approval as they are currently doing tests that are available on commercially and FDA approved devices already (this makes sense because LDTs were never ment to be used in place of standard tests and equipment and since there is NO published data on their performance it can be worrisome to providers and payer)
Their true business plan has yet to be elucidated, but if they really did have a box that did all this, they would be selling them to physicians and clincs where those results could actually be used in diagnosis rather than 4 hours later.
I hope they succeed, but there is just way too much hype to buy in on it yet.

I am sorry I don't have sources, most of this is from intimate knowledge of the space and from the little info available on them so I could definitely be wrong on some of this.  I hope this helps!

Thank you very much for your excellent answer with outstanding details and analysis! I am interested in in the Biological and Physical limitations here, particularly. They do not cover anywhere the uncertainties of their tests. I would like to work for a review about uncertainties in sampling small volumes of blood in different tests. How did you find the pieces of information about their patents? Google patents with Theranos gives 10 international classifications: A61B5/00, A61M5/172, B01J19/00, A61B5/15, B01L3/00, A61M5/142, A61M37/00, A61M5/145, G01N35/00 which I think are their patents

Issued Patents:https://www.google.com/?tbm=pts&gws_rd=ssl#q=inassignee:%22Theranos,+Inc.%22&tbs=ptso:us,ptss:g&tbm=pts  Applications:https://www.google.com/?tbm=pts&gws_rd=ssl#q=inassignee:%22Theranos,+Inc.%22&tbs=ptso:us,ptss:a&tbm=pts&start=10

Wow! This is insightful answer, clear expert on this area! I tried to summarise this answer in my answer related to sensitivity and amplification. I cannot fully understand them. Perhaps you could preview them? +1 for the awesome work already!

I haven't seen their device and I won't want to see it, I want to first understand the concept better independently. I am not an expert in biology but I have feeling how such device could be done.
Noname covers sensitivity and new concept "amplification" raising a new puzzle: is amplification done by analog methods (op-apms), digital methods, dilution or/and some other methods?

Summarising the Noname to sensitivity, amplification and background

(I added bolding)

"Theranos is working on modular micofluidic devices/cartridges"

"regardless of how good your device is, at some point you start amplifying junk. The better the primers and other reagents the less junk you amplify, but at the end of the day you want more sample to be more sensitive."

"they aren't/can't multiply the analyte of interest. In these tests, they rely more on dilution or small volumes to perform detection. This is where their ability to reliably dilute and correct for the dilution comes into play. -- if you had perfect detection you might be ok, but you generally want to analyze more blood so that statistics work in your favor and your performance goes up. Most of the other chemistries, proteins, and as mentioned earlier, electrolytes will probably be ok for dilution and detection and in fact this is why there are plenty of small devices already out there that use a drop of blood and perform the same tests."

"They don't publicize that they may need to draw a venous sample (needle in your arm) for some tests and depending on what tests you order, you may need a separate finger prick for each test."

where by (2) more sensitivity can actually be a good thing; by (3) same tests already done by other devices in small size: what is so special with Theranos device? and by (4) one drop of blood not enough for all tests.
"What are limiations of multiplying so little blood?"

Decision Analysis (DA) and Compositional Blood Analysis (CBA): I see such a device as a Decision Analysis (DA) device where you need to do compositional blood analysis (CBA). You get a lot of vague information on which you do sensitivity analysis. Conventional full blood tests can be seen as devices where the sensitivity is very very low to other factors. It is not surprising that development has lasted about 10 years, sensitivity analysis is very low research project. For example, Chevron's DA model started 1990 and continuing, implementation takes time. For example, scale hydration level to some level in all samples. When you have low-quality information, you need to use ad hoc models (often based on high-quality samples) such as supposed correlations between things to improve prediction accuracy.

1.1. What are Decision factors?

1.1.1. Rheology of blood? Dry or liquid phase samples?
1.1.2. Correlation of red/white shells: I don't use the word "approximation" because the number of red shells correlated with the number of white shells
1.1.3. Density (~1.05 kg/l)
1.1.4. Colour of blood
1.1.5. Hydration level
1.1.6. other factors?

"How can Theranos blood test's be specific and accurate compared to conventional full blood test?"

I think this question mis-framed. It is better to ask "How much extra value does such a device can offer?"
Smaller size and less blood mean convenience. It may be useful with athletes, new markets. More convenient to patients/users even though suffering from some abnormalities due to its intrinsic limitations related to its small-volume measurement method.

What components do you need to multiply?

I think this is an important insight: is it possible to multiply blood somehow ensuring its quality for testing? This would make blood tests less painful for patients. Perhaps this is related to stem-cell research? Perhaps not, the Noname covers this in terms of dilution more closely.

Using ">" indicates that you are citing a source. Are you? If so please provide a link. Otherwise please format your post differently.

Please, see Noname's answer. I think you need to revise your answer accordingly.

@Masi Christian: done done, ">" changed to "---" and the Noname summarised related to sensitivity and amplification. I think the Noname adds more insight to sensitivity on this process. The DA and CBA are still the underlying processes in building the product -- where I think most other small-size products fail (it requires capital/time/creativity/teamwork/etc).

-1 I think your answer is too much business-minded: Extra value of device. Specificity and accuracy of the device in some test is very relevant here. I think you are little bit lost in your last big title question - PCR is for multiplying  blood parts. Those patents seem to apply too it and to multiply just some parts of blood - see Noname's answer. Rheology of blood should be able to maintained by pharmacological methods and by method of educating personnel who take the blood. So no coagulation problems. I think your listed factors are not keys here. Those "other factors" are most relevant.


text:
Can the human eye distinguish colors in the periphery?
In the back of my mind I have the idea that human eyes can't notice the color of objects in the far periphery, and that any subjective perception of colors is done by the brain that tries to fill in the color percept.
Is that true?

It depends to some degree on what you define as peripheral vision. Until relatively recently, color vision in the peripheral field has been thought to be substantially less developed than color vision in the central field.
Most estimates of peripheral color perception place the limit of trichromatic (RBG) vision at no more than 30 degrees from fixation; the visual field was thought to be completely color blind at about 50 degrees of eccentricity.
Within the last 30 years, an increased understanding of the changing spatial scale in the peripheral field has led researchers to reevaluate what is believed about peripheral function. Virtually every measure of peripheral color perception can be improved by using a suitably large stimulus in the peripheral field.1
Here is a schematic of distribution of cones in the retina. While cones are decreased in number at 60°, they do exist.2

Here is a schematic of angles on peripheral vision.2

Previous measured sensitivity curves farther into the periphery than thought. In one paper, detection thresholds for colored disks showed that color vision, in particular color vision mediated by the L – M cone-opponent channel (luminance or blue–yellow colors), persists up to at least 50 deg. The larger the color disc, the better the peripheral color vision.[3]

The range of eccentricities over which red–green color vision is still possible is larger than previously thought. Color stimuli can be reliably detected and identified by chromatically opponent mechanisms even at 50 deg eccentricity. Earlier studies most probably underestimated this range. Differences could be caused by technical limitations and the use of stimuli of non-optimal size. (Emphasis mine) In agreement with previous studies we found that the decline in reddish-greenish L − M color sensitivity was greater than for luminance and bluish-yellowish S − (L + M) signals. We interpret our findings as being consistent with a functional bias in the wiring of cone inputs to ganglion cells (Buzás et al., 2006) that predicts a decrease but not a lack of cone-opponent responses in the retinal periphery.

1 Color vision in the peripheral retina
2 The Retina: Rods and Cones
3 Color perception in the intermediate periphery of the visual field

There are two types of photoreceptor in the eye, known as rods and cones. Rods are spread equally throughout the whole retina, whereas cones are focussed in one spot known as the fovea:

Cones are the only type of photoreceptor that can sense colour, so rods can only sense black and white. However, rods can work in much lower light and will also work when not being focussed on. Due to what is known as 'summation of action potentials', through which nerve impulses from multiple rods add together to cause a signal to be sent to the brain, rods are much more effective than cones at sending impulses. If you've ever seen something flickering in the corner of your eye and not been able to see what it is when you've looked properly, this is probably why (Wikipedia: Peripheral vision characteristics).
Edit: Further information on how vision is perceived can be found in this answer.
As for the question of whether or not the brain tries to guess the colour I don't know. I would suggest yes, the brain compensates as much as possible for a lack of information. It's known that the brain can compensate for the blind spot where the optic nerve passes out of the eye and compensates for it using the other eye and other information (Why every human has a blind spot, and how to find yours). However that is just my opinion, I haven't been able to find any research relating to the idea.


text:
How can Pyridostigmine have the indication of Myasthenia?
I am thinking how pyridostigmine can be used in the treatment of myasthenia gravis.
Its similar compound (neostigmine) is also anticholine esterase.
This compound has the indication of myasthenia gravis but with the stimulation of the nicotinic receptor.
I cannot understand how pyridostigmine is causing the effect.
Probably, pyridostigmine is similar to neostigmine in structure, which can be why more pieces of information is about neostigmine than pyridostigmine.
How can pyridostigmine have the indication of myasthenia gravis?

Pyridostigmine is an analog of neostigmine, so has the same basic mechanism of action. Please forgive me for repeating information you may already know.
There are two main families of acetylcholine receptors: muscarinic and nicotinic. Acetylcholine receptors at the neuromuscular junction (NMJ) are exclusively nicotinic (N1 or NM), so that's the only receptor I'll address.
Myasthenia Gravis is caused by an immune response to nicotinic postsynaptic receptors on skeletal muscle. An antibody can either destroy or block the receptor site. The antibodies destroy the receptor sites more rapidly than the body can replace them. In MG, there can be as much as an 80% reduction in the number of these receptor sites.
Acetylcholinesterase (AChE) is an enzyme that breaks down acetylcholine (ACh) at the synaptic cleft. AChE has a remarkably high specific catalytic activity: each molecule of AChE degrades about 25000 molecules of ACh per second. Any drug which serves to inhibit AChE will increase ACh concentration at the N1 receptor. AChE inhibitors can be irreversible or reversible. Reversible inhibitors, (competitive or noncompetitive) have therapeutic applications; irreversible AChE  modulators are toxic.
Carbamates are organic compounds derived from carbamic acid (NH2COOH). They are reversible AChE inhibitors. This group includes neostigmine, pyridostigmine and physostigmine. Pyridostigmine is an analog of neostigmine, so has the same basic mechanism of action. Because they are not lipid soluble, they are selective for the NMJ; both also have direct action on Nm Receptors. The major difference between them is that pyridostigmine has a slight but significantly longer duration of action.
Drug Bank: Pyridostigmine
NeuroLex.org (get's it's info from PubChem)

Thank you for your excellent answer with great details! It is never wrong to repeat. Pyridostigmine is parasympathomimetic  and reversible cholinesterase inhibitor. These two features seem to work in different directions, but the latter is the main one. It leads to the property of reversing muscle relaxants and its indication of myasthenia gravis. The former effect does not seem to be a major one. -- Let's consider Pilocarpine which is cholinomimetic and exclusively muscarinic (nothing about nicotinic?). It seems that only acetylcholine inhibitors are exclusively nicotinic with (N1 or Nm).

If I'm understanding you correctly (and I'm not sure that I am) pilocarpine is a muscarinic agonist with no nicotinic effects. But there are AChE that are both Nm and muscarinic.

You are understanding me correctly. It seems that also the other short-acting and long-acting AChE are Nm and nicotinic besides neostigmine and pyridostigmine such as ipidacrine, donepezil and rivastigmine.


text:
Purifying a linear plasmid after restriction digest?
I expressed a yeast vector in E.coli and purified about 13µg of it. I then linearized it using a restriction enzyme, and attempted to gel purify it. I attempted this twice. The gel showed a clear bright band at the right spot, but after gel purification I end up with only 0.7µg of linear plasmid. I doubt I made the same mistake twice. What is the best way to purify a linear plasmid after digest?
Edit:
The plasmid is about 3.5 kb. I incubate an overnight culture of E. coli, miniprep the following morning, elute into water. I end up with 13µg of DNA in 200µl of H2O. I add the PmeI (10µl) and the CutSmart buffer (23µl) from NEB, incubate overnight. The following day, I run the 230µl through a 1% gel and get a single band around 3.5 kb. I excise the band, and perform a gel purification using the Biobasic kit, add denaturant (probably guanidine hydrochloride), wait for 10 minutes for the gel to dissolve, run through column, wash 2X with wash buffer with ethanol added, elute in water.
Thanks,
Max

Could you share the size of your bands with us and give a bit more details about your current purification method!

The plasmid is about 3,500 kb. I incubate an overnight culture of E. coli, miniprep the following morning, elute into water. I end up with 13 ug of DNA in 200 uL of H2O. I add the Pme I (10uL) and the CutSmart buffer (23 uL) from NEB, incubate overnight. The following day, I run the 230 uL through a 1% gel and get a single band around 3,500. I excise the band, and perform a gel purification using the Biobasic kit, add denaturant (probably guanidine hydrochloride), wait for 10 minutes for the gel to dissolve, run through column, wash 2X with wash buffer with ethanol added, elute in water.

230uL of solution seems like a lot to put on a gel. Are you using multiple lanes and therefore putting a lot of gel through the gel extraction kit? I know they can only handle so much gel at once.

Could you please make this as an edit to your question! Why do a gel purification? you can just use a kit similar to QIAGEN gel extraction kit and do a non gel based plasmid purification and elution, using the buffers used as usual! PCR DNA clean up kits will do as well!

Good idea. I will attempt to purify using PCR cleanup. Thanks Bez!

What is the loading capacity of the columns you used for the purification?

Which kit did you use exactly?

@Max if that has answered your question please let me know so that I can post the response as an answer.

Yes, please do!

I have a quick question: How are you growing your culture? Is it a liquid culture? The reason I am asking is that you DNA concentrations sound very low before you cut the DNA. I regularly grow E. coli in a 4mL LB in H2O for subsequent DNA purification using the QIAGEN plasmid mini-prep kit (3mL protocol) and I get nearly 120ug/uL of DNA with A60/A280 of ~1.9.

Hello gravityassist,
I grow about 20 ml overnight in low-salt LB with Zeocin (25 mg/ml). I do 4 minipreps and end up with 200 uL of 65 ng/uL. We tend to get 50ul of between 50 - 120 ng/ul for our low-copy plasmids.

3500kb???!!!.. Are you sure? I think it must be 3500bp=3.5kb.

3,500 bp. My bad.

Can't exactly say what problem you are facing but you can follow these steps for efficient restriction digestion and elution.

Never set up a digestion reaction with more than 1µg DNA per 20µl; if you require more digested DNA, then set up multiple 20µl reactions with maximum of 1µg DNA in each (500-700ng is ideal).
10µl enzyme for a 230µl reaction: bad idea. Set up 10×20µl reactions with not more than 0.5µl enzyme per reaction tube.
Avoid overnight incubations; 4 hours should be sufficient (NEB PmeI is 10 units/µl and 1 unit is the amount required to cleave 1µg DNA in 1h in a 50µl reaction. You are adding 5 units in 20µl. Overnight incubation is too much)
Deactivate the enzyme after the reaction.
Just run a little bit of sample from one tube in the gel to see if digestion has happened. After confirming digestion simply use the PCR purification kit. You can pool your tubes.
Optional: Elute in Tris-Cl pH 7.5 instead of water or use warm (60⁰C) water.

Additional: Check the maximum DNA capacity of the columns used. If you overload them, you will drastically loose yield.

@Chris That is an important and nonintuitive problem, I once wasted a whole gigaprep kit because the protocol a former lab member gave me said to use twice as much DNA as the column could hold, then my advisor thought I could fix the low yield by loading even more DNA.

@user137 I have run into this problem as well, this is why I mention it.

@user137 note that a column can be reused. So you can use it for multiple rounds of elution for same plasmid.

@WYSIWYG How should I clean up the column to reuse it? Gigaprep kit are expensive and we only produce 1 plasmid on that scale so reusing a column should be ok for us.

@user137 well you can add a little bit of tris-Cl pH 7.5, spin it for 2min at 12000rpm for 2 min to remove residual stuff and then reuse.

@WYSIWYG We may be confusing gigaprep and miniprep, the Qiagen gigaprep column looks about 6 inches long with diameter of about 2 inches and works by gravity. Even if it can survive centrifugation I don't have a rotor that large.

@user137 ahh.. then you would probably need vacuum pumps.. Nonetheless for the same plasmid type it is not an issue. you can reuse it without much worry. just use vacuum before reuse to ensure that nothing stays in the column.

Why do a gel purification? you can just use a kit similar to QIAGEN gel extraction kit and do a non gel based plasmid purification and elution, using the buffers used as usual! PCR DNA clean up kits will do as well!


text:
Where should I start looking for an online gene library?
Is there an online library that includes all known genes in stored in their nucleotide form of all living organisms?
I'm presuming that such a database doesn't exist. So to kick of my search, how can I find a gene amongst all the other databases? Where is a good place to start looking for information about my gene given that it is a well studied sequence? The issue is that I have been provided with the sequence, but I just don't know what organism the sequence belongs to or what it codes for.

Databases
There are too many databases to list here, but Wikipedia has a decent list of the genome databases. For example, there are databases dedicated to genes of individual species, like Wormbase, SGDB and countless others. Alternatively, as others mention a great place to kick off your search is the NCBI.
Finding your nucleotide sequence in databases
It sounds like you already have your sequence. If you want to see if your sequence has been found elsewhere or if there are any similar sequences, you can use BLAST from NCBI. Although it is not strictly a database in itself, it looks to align your sequence to similar sequences in external NCBI databases and quantifies the sequence similarity. It will reveal the species for you to go to a more appropriate database along with a lot of other information about the gene.
There are plenty of parameters to alter to suit your specific needs, but generally, the default settings are a great place to start analysing your gene. You'll need to run a nucleotide test since you want to find the DNA sequence. The results might be difficult to interpret.

NCBI is a good option.
http://www.ncbi.nlm.nih.gov
If you're looking for a gene sequence,  you can search for the gene name in the nucleotide database. For example, here is the annotated human insulin gene;
http://www.ncbi.nlm.nih.gov/nuccore/J00265.1

SwissProt is a human-annotated database of genes/proteins with known functions.

Although this does have the gene information (or at least information on where to access information on the genes) this is a protein centric database and deals in amino acid rather than nucleotide sequences.

There is no properly annotated DNA database. Use this to find function of interest and use a tBlastn to find the gene. Alternately, BRENDA is a human-annotated enzyme database. Use tBlastn to find gene.

No, you are wrong in your assumption that there is no such library.
GenBank. One-stop shop.
Blast your sequence against it on-line. If it isn’t there you are not likely to find it.


text:
Is there a stem cell dye for education purposes?
I need a dye that will stain stem cells (for this purpose the stem cells will proquired from chicken bone marrow).
This will be an educational hands on activity for young teenagers so ideally the dye should be:

Safe
Easy to handle
Stain quickly
Be relatively cheap
Present an obvious colour change under a low zoom microscope or the naked eye

I vaguely remember using a dye years ago that fits this bill, but cannot recall the name or company.

There is no specific dye for stem cells. You would have to do an immuno-histochemical staining for stem cell markers such as Sox2/Oct4 etc.
Usually stem cells have a distinct morphology (round and clustered). You can use Leishman's (or Romanowsky-Giemsa) stain.

How about Brdu...? Normally, stem cells have active proliferation, their dividing are fast, thus they could uptake Brdu, and this is one of basic stem cell test in papers. I think it isn't too expensive, comparing with antibodies.... but you may need UV-light, and don't let children play Brdu. It is carcinogen, however, I think it only a very high dose can cause a cancer...