Antibody Quantitation Using an Immunoabsorbent and the Unlabeled Antibody Enzyme Method.
Technical rept. May 72-Jul 74,
EDGEWOOD ARSENAL ABERDEEN PROVING GROUND MD
Pagination or Media Count:
The measurement of specific immunoabsorbent-bound antibodies has been amplified by the unlabeled antibody enzyme method J. Histochem. Cytochem. 18, 315, 1970. Sepharose-4B containing specific antigen or ligand is treated with diluted specific immune serum primary serum, such as 1 ml of serum diluted 6000-fold to 20,000-fold, followed by antiserum against immunoglobulin G IgG of the primary serum and by the peroxidase-antiperoxidase antigen-antibody complex PAP derived from the same species as the primary serum. Radiolabeled primary antibody and anti-IgG have confirmed and stoichiometry of the reaction. The immunoabsorbent binds the antibody of interest quantitatively and to equal extent after 15 minutes or 48 hours. The enzymatic activity of the PAP complex followed a direct linear relationship to its concentration indicating the stability of binding in the PAP complex. A direct relation between the enzymatic activity measured for both the primary antiserum and the anti-IgG allows for quantitation of the antibody level of the primary serum. The effective stoichiometry of the system was 111 -specific antibodyanti-IgGPAP.
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