Protein antigens molecular size

Elute the antigen ( MAb) by heating beads (5 min at 95°C) with equal volume of SDS-sample buffer, and apply to an SDS-containing polyacrylamide gel. Run prestained markers on these gels, because they will transfer to blots and assist in determining the molecular size of the proteins. 

Cmde enzyme extracts are often unsuitable for therapeutic uses because of their antigenicity, contamination with endotoxins, and rapid inactivation under physiological conditions or in fluids intended for intravenous infusion over several hours. When the enzyme used is a foreign protein, it can eHcit an immune response that alters the clearance rate or induces severe allergic reactions in the host. After an intravenous injection of an enzyme, its activity in plasma decreases with time due to distribution to other fluids and tissues, and as a consequence of proteolysis or excretion. Distribution is related to molecular size, charge, and HpophiHcity surface charges attributable to the availability of free amino, amido, or carboxyl groups may affect the rate of inactivation of some enzymes. 

Now, while it is possible, by lowering the surface tension of the liquid medium to 50 dyn/cm by means of solutes of low molecular weight, to dissociate antigen-antibody precipitates (of the pure VDW type) (6), whole mammalian serum (with a surface tension < 50 dyn/cm) definitely has no such dissociating power. Upon some reflection this is not as anomalous as it might seem. Table I shows that it is mainly because of the presence of proteins (of molecular weight > 20,000) that blood (or plasma) has a surface tension y < 50 (and not y 70), and it cannot be expected that molecules with dimensions of the order of 100 A will effect a separation by purely physical means between other proteins of the same approximate size that are only about 2 A apart at their site of interaction (13). 

For many proteins, a purified protein standard is ideal and straightforward to incorporate since the concentration is known. However, in reality such standard/ sample accord is often not possible to obtain because the antigen contained in the sample may be a mixture of species. For example, it may come in a variety of molecular sizes (e.g. monomeric versus oligomeric forms of immimoglobulin A), or it may exist in complexes with other molecules (e.g. a protease bound to a macromolecular inhibitor, or a ligand bound to its soluble receptor), or as a... 

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