Enzyme immunoassay for the quantitation of human immunoglobulin G specific for the glycolipid of Enterobacteriaceae.
An enzyme immunoassay (EIA) is reported that quantitates human immunoglobulin (Ig)G specific for the glycolipid (Re-GL) of Salmonella minnesota Re 595. Re-GL was adsorbed to polystyrene microcuvettes, and 100 microliters of appropriately diluted lots of human plasma or affinity-purified rabbit anti-Re-Gl IgG was added to the cuvettes. The cuvettes were incubated, washed, and then reacted with Staphylococcus aureus protein A-peroxidase. After a final wash the substrate 2,2'-azino-di(3-ethylbenzthiazoline)sulfonic acid (ABTS) was added to yield a blue-green color with a maximum absorption at 405 nm. Using the affinity-purified rabbit anti-Re-IgG as standard, it was possible to quantitate specific levels of anti-Re-GL. The EIA was rapid (2 hr), with an absolute sensitivity of 5 ng of specific anti-Re-GL IgG and an interassay coefficient of variation of 24%. On a unit-mass basis, rabbit anti-Re-GL IgG had 10-fold greater activity than comparable affinity-purified human anti-Re-GL IgG from normal donor plasma. When a rabbit standard was used, the mean level of specific anti-Re-GL IgG in lots of pooled human plasma, which consisted of 18,330 liters, was 1.1 +/- 0.28 micrograms/ml. Of 237 individual donors tested, 24 (10%) had levels of anti-Re-GL IgG that were 10-fold or greater than the mean of 1.1 micrograms/ml. The EIA provided a rapid, reproducible method for accurate quantitation of specific anti-Re-GL IgG in the screening of large volumes of human plasma.
D000818:Animals; D000907:Antibodies, Bacterial / Q000032:analysis*; D000918:Antibody Specificity; D004755:Enterobacteriaceae / Q000276:immunology*; D006017:Glycolipids / Q000276:immunology*; D006801:Humans; D007124:Immunoenzyme Techniques; D007074:Immunoglobulin G / Q000032:analysis*; D011817:Rabbits
D000907:Antibodies, Bacterial; D006017:Glycolipids; D007074:Immunoglobulin G