Coupling of different isoenzymes of horseradish peroxidase influences the sensitivity of enzyme immunoassay.
Labelling of IgG with various HRP isoenzymes purified by preparative isoelectric focussing influences the yield and the specific activity of the conjugates. Alkaline isoenzymes were preferably coupled by glutaraldehyde whereas application of the periodate method additionally formed relatively large quantities of conjugates with acidic isoenzymes of a high purity number, but a low specific activity. In an enzyme immunoassay for alpha fetoprotein the detection limit can be varied by a factor of 6 and even by a factor of 20 by use of the conjugates with different isoenzymes coupled by the glutaraldehyde method and the periodate method, respectively. In order to achieve enzyme immunoassays of the highest sensitivity, antibodies should be coupled to horseradish peroxidase after removing acidic isoenzymes from the enzyme preparations.
D000918:Antibody Specificity; D005976:Glutaral / Q000032:analysis; D006735:Horseradish Peroxidase; D007124:Immunoenzyme Techniques; D007074:Immunoglobulin G / Q000032:analysis; D007525:Isoelectric Focusing; D007527:Isoenzymes / Q000276:immunology; D010504:Periodic Acid / Q000032:analysis; D010544:Peroxidases; D000509:alpha-Fetoproteins / Q000032:analysis
D007074:Immunoglobulin G; D007527:Isoenzymes; D000509:alpha-Fetoproteins; D010504:Periodic Acid; D006735:Horseradish Peroxidase; D010544:Peroxidases; D005976:Glutaral