Quantification of immunoglobulin on electrophoretic immunoblot strips as a tool for human immunodeficiency virus serodiagnosis

Abstract

Electrophoretic immunoblotting (EIB [Western blotting]), the main method for verification of human immunodeficiency virus (HIV) seropositivity, needs thorough characterization and standardization. We explored the possibilities of quantifying immunoglobulin G (IgG) bound to EIB strips both by densitometry of the peroxidase-stained bands and by measurement of radioactivity with labeled anti-HIV IgG. The radioactivity method is inherently more exact but was more cumbersome. However, despite saturation phenomena at high IgG densities, the densitometric method was more convenient and yielded reproducible estimates of the amount of bound IgG. We found it useful primarily for documentation of changes in the relative abundance of antibodies to different HIV proteins from individual patients over time. To explore the potential usefulness of the method, we studied a small set of HIV-seropositive persons. The average p24/gp41 color yield ratios and standard deviations in 3 persons with recent seroconversion, 15 healthy subjects, and 6 diseased HIV-seropositive persons were 6.6 +/- 0.9, 2.3 +/- 1.9, and 1.3 +/- 0.5, respectively. These data are in accord with previous qualitative or semiquantitative observations but are too limited for any conclusions regarding the use of quantitative EIB for prognostic use with individual patients. Quantitative EIB is a valuable tool for comparative methodological studies and for research on the protective role of anti-HIV antibodies in acquired immunodeficiency syndrome pathogenesis. Its possible use in prognostication for individual patients must be evaluated in long-term studies.