Preparation and standardization of perotidase-labeled anti-human IgG antibody for use in determination of serum antinuclear antibody levels.
Peroxidase-labeled antibody conjugates were prepared by a two-step conjugation procedure using glutaraldehyde. Immunoadsorbent-purified antibody and the gamma-globulin fraction of sheep anti-human IgG antiserum were employed for these preparations, Procedures for the determination of the antibody and enzymatic activity, as well as the specificity of the enzy-e-antibody reactions, were outlined. Peroxidase-anti-human-IgG conjugates were prepared with approximately a 1:1 molar ratio of peroxidase to IgG, which demonstrated a loss of 10-15% of precipitating antibody activity. Standardization procedures for use of peroxidase-labeled antibody in the indirect ANA test were established. The peroxidase-labeled antibody in the indirect ANA test were established. The peroxidase-antibody procedure was found to demonstrate a reproducible plateau endpoint which was comparable to that of the fluorescent antibody preparations.
D000818:Animals; D000906:Antibodies; D000974:Antibodies, Antinuclear / Q000032:analysis*; D005455:Fluorescent Antibody Technique; D005976:Glutaral / Q000378:metabolism; D006801:Humans; D007106:Immune Sera; D007074:Immunoglobulin G / Q000302:isolation & purification / Q000378:metabolism; D007158:Immunologic Techniques; D010544:Peroxidases / Q000378:metabolism; D011817:Rabbits / Q000276:immunology; D012756:Sheep / Q000276:immunology; D005719:gamma-Globulins / Q000302:isolation & purification
D000906:Antibodies; D000974:Antibodies, Antinuclear; D007106:Immune Sera; D007074:Immunoglobulin G; D005719:gamma-Globulins; D010544:Peroxidases; D005976:Glutaral