When using antibodies to make affinity chromatography, which molecules used to fix the antibody to the column?
607444
...                                                                               \n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n                          You want to use the "best" molecules.\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n\n                                                       .
1057303
do you mean monoclonal antibodies?  If yes - it can be given IP in mice - never seen it used in rats before.  You would need to follow the SOP's at your facility - it would explain the proper procedure. I have never seen mini pumps used for monoclonal antibodies.\nDo you not need protocol approval first - check with the techs/vets at your facility.
431813
Tag it with a poly ubiquitin chain.  You can make a chimera of the protein plus a couple of ubiquitins.  We did this for our protein (one ubiquitin tag at C terminus) and found it was degraded very efficiently.
353450
Aantibodies when they bind to the corresponding antigen(s) bring as many as 5 such antigens or antigen containing entities per antibody depending on the type of the antibodytogether. However such antigen containing cells/entities have many such antigens on their surface and therefore are attacked upon by many such antibodies which result in a meshwork like formation resulting in the phenomen of agglutination. The classic example for this is blood grouping.
1021546
Let's see if this helps...affinity is merely the ability for antigen to bind an antibody...it can however be reversed...so it isn't extremely affective. Affinity describes the strength of interaction between a single paratope(antibody) and a single antigen. Avidity refers to the strength of interaction between a multivalent antibody and antigen. For example...an antibody fragment only has affinity and isn't very affective. Also, if IgG only has 1 receptor bound by antigen...it only exhibits affinity. Now, if both receptors of IgG are bound, it become avidity. Or if the pentamer of IgM is bound at 2 or more receptors...it is also avidity. It is a very strong attraction that will more than likely not be broken.
116691
Chromatographyis a way to seperate out different parts of a mixture.  The idea is that you pass a fluid (either a gas or a liquid) down a long path, and the different molecules will move at different rates.  THat's the long and short of it.  So the molecules that move fastest get to the end first, and the slowest come out last.  You seperate things by making them race, and similar chemicals will take a similar amount of time to finish the race.\n\nI'm assuming that you're getting confused by things like "solid phase" and "mobile phase" and "bind affinity" and such.  The idea though is that you set up different types of obstacles depending on which things you want to win the race.  Sometimes this is dine by using something with a lot of positive charges, so that things with negative charges will get stuck and the things that are neutral or positively charged will win the race. Sometimes you sort things by size, by using a gel as an obstable with a bunch of little pores in it: little guys get lost inside and the fat guy wins the race.  With gases, you can just make the race really, really, really long, and the lightest parts will move the quickest while the hevy boys lag behind (this is a way that uranium is enriched for nuclear reactors and bombs).
1125782
So you need chlorobutane to run a column?\n\nWhat about something similar, in terms of polarity, like chloropropane or chloropentane?\n\nI was going to run a column once based on results from TLC using a pentane/acetone mixture. The lab ran out of pentane and the stock room was closed. My undergraduate research advisor suggested hexane instead. We tried it, it worked, and I was able to get the separation.
919511
Chromatography is used mainly to quantify or measure purity and the concentrations of impurities present in a sample.\n\nTLC is used to used in many Quality Laboratories to measure the impurities present in raw materials.  Most amino acids, along with some other active pharmaceutical ingredients, used in the pharmaceutical/biotechnology industry require impurity testing by TLC.
621891
One is a simple binding assay, in which the ligand (usually radiolabeled) is added to a hemogenate of the tissue-a number of washes are done, in the presence of cold ligand, in order to retain only that ligand that has bound with high affinity.\nThe others are all protein-based.  Western blotting with specific antibodies, immunohistochemistry to determine the presence and cellular localization.  RT-PCR or Northern blotting will tell you if the mRNA is present, but not if the receptor protein itself it there.
689265
Antibody molecules are Y shaped structures.\n\nThe two upper ends of Y has got the sequences for the binding with the antigen and the lower end has constant amino acid residues.Also they got 2 chains called the heavy and light chains.\nThese sequences are coded by the genes of the B cell.\nThe antibodies are classified based on their heavy chains into igG,IgM,IgA,IgE,IgD and antibodies within such a group will have their constant site(lower end) resembling each other and only the variable regions would differ(upper ends).\nThus the virtual identity regions must be the constant region I said.
115334
Generally polyclonal anitbodies are used to bind non-specifically to antigens. For instance, you might use monoclonal antibodies to specifically bind to your protein of intererst. You might then use polyclonal Ab to bind the rest of the proteins i.e. to block off the other available 'slots'
973006
okay I'm gonna try this. :-)\n\nImmunohistochemistry is a way to detect certain proteins that are in your body's tissues.\n\nIn kidney diseases, there may be abnormal proteins in the kidney that you'll want to look for.\n\nSo what happens is a scientific technician will take an extremely thin slice of kidney tissue, and treat it with "antibodies" to the abnormal (bad) protein.  \n\n[An antibody is something that will stick to a certain protein, they can be made to be very selective.  So there is a specially designed antibody that will find this bad protein and stick to it.]\n\nIn addition, this antibody has a "signal" stuck to it.  Normally it's some kind of fluorescent (shiny, hehe) dye.  After treating the sample with antibodies and getting them stuck to the abnormal proteins in the tissue, the technician will then perform a reaction that will activate the dye.  Then, the sample is put under a microscope, and the picture is analyzed for presence of this dye.  If the kidney is diseased and has the abnormal "bad" protein, there will be colored dye spots all over the picture.\n\nSo in short... kidney diseases can show abnormal (bad) proteins, scientists can make things ("antibodies") that stick to these bad proteins, and then these antibodies can be "activated" to light up the dye and reveal where the bad proteins (which they are sticking to!) are.\n\nIn theory, a normal kidney tissue sample would not show any fluorescence/lighting if they were looking for a "bad" protein.  Of course, "bad" protein is a huge generalization, and I'm just trying to simplify it into layman's terms. \n\nContact me if you have other questions, hope this helps!\n\n-IJ
313806
Affinity purification is a method commonly used in protein chemistry to isolate a product from a mixture that interacts with another known, and previously isolated, product.  Negative affinity purification, on the other hand, is a method used to deplete a known contaminant from a protein mixture based upon its ability to bind to a different molecule.\n\nFor example, a scientist wants to make an antibody specific for a novel protein. He/she produces the protein as a fusion product linked to GST (glutathione-S-transferase), since there are many methods to allow isolation of this fusion protein (by affinity purification techniques).  This fusion product is then injected into a mouse or rabbit and after some time the animal produces antibody in its blood.  \n\nWhile it might be acceptable to use the whole serum (containing the protein specific antibody) from this animal for subsequent experiments, the antibody of interest is only a small fraction of the total protein in the serum.  So, the scientist purifies the antibody directed against the protein of interest using a column containing the GST-fusion product used for inoculation (again, affinity purification) and elutes the antibody fraction that recognizes this protein.  Great, but the scientist knows that the GST component of the fusion protein will also induce the mouse to make antibodies, and that these GST-specific antibodies will be purified by the previous step.  So, he/she decides to do NEGATIVE Affinity Purification by passing the purified antibody mix over a column that contains the GST protein alone (not the fusion product) and then collects the antibody that DOES NOT bind to the GST alone.  Voila, protein specific antibody without contaminating GST-specific antibody... using negative affinity purification.
900028
Compounds injected onto a column interact with the column material. Some compounds "stick" to the column more than others. These compounds then have a long retention time. Heating increases the kinetic energy of the compounds on the column. This increased energy allows the molecules to wiggle free from the column more easily. So heating reduces the "stickiness" of the molecules. Since molecules will "stick" less, they will move more quickly through the column, and their retention times will decrease.
42016
A modification of immunoelectrophoresis in which antigen and antibody move in opposite directions and form precipitates in the area between the cells where they meet in concentrations of optimal proportions.\n\nHelped?;)
816465
Two substances must flow through the medium at different rates. High performance liquid chromatography operates by running a liquid through a solid column. The molecules with the same chemical properties as the solid will take longer to go through the column. If the solid is polar and the liquid is non-polar, then non-polar molecules will move through faster because they do not get stuck in the solid. Bead chromatography involves running a liquid through a column loaded with gel beads that contain some holes. Smaller molecules spend some time in the holes, but molecules too large to enter the holes go straight past the beads and exit the column first. There are different ways to separate substances and there are a ton of different chromatography methods.
116211
Probably you mean "rabbit anti-goat IgG" antibodies, which are commonly used in biomedical reasearch labs. It works as follows: let's say for your science experiment you need an antibody against a certain protein (let's name this protein X). You inject X into a goat and over time the immune system of the goat generates antibodies against X (let's call these antobodies goat anti-X). Antibodies of type IgG are the most abundant at the latest stage of immune response and we isolate them for our experiment (so we obtain goat anti-X IgG). For certain purposes (I won't go into details) we may need another set of antobodies which would be against our goat anti-X  IgG we just got. How do we get them? We cannot generate them in another goat, because the immune system of the same species is unlikely to respond to its own product. Therefore, we inject another species (for example, a rabbit) with our goat anti-X IgG. The immune system of the rabbit recognizes the foreign molecule and generates an antibody against it. It will be the rabbit anti-goat IgG - it means, this rabbit antibody will recognize the previous antibody for the goat. We then can further use it in our experiment.