For antibodies provided in a serum format (polyclonal) or not purified supernatant (monoclonal), the final concentration of specific antibody is not estimated. Instead of specific antibody concentration, one has to use recommended dilution of an antibody in a given technique as a guideline. Following antibody purification, also called affinity purification (on Protein A or G), a total immunoglobulin fraction is obtained. The concentration of specific anti-target antibodies is not determined. It usually varies from 0.5 to 5 % of the total immunoglobulin pool. Specific antibody concentration can be estimated following affinity purification on an antigen (peptide or protein), coupled to the appropriate matrix. In such cases, the concentration of the specific antibody is provided on the tube, or can be calculated following reconstitution of a known amount of purified antibody. For example, 50 µg in 50 µl of deionized water will give specific antibody concentration of 1 µg/µl. |
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