Determination of theoptimum Sample dilution of Commercial TotalIgGELISA, the Total and anti-measles immunoglobulin G in HIV infected pregnant women

Abstract

Enzyme-linked immunosorbent assays (ELISAs) are widely used to quantify immunoglobulin levels induced by infection or vaccination. The first large-scale use of ELISA for antibody quantitation was in diagnosis and epidemiological viral screening to assess antibody levels after vaccination. Measles virus is transmitted through infected air droplets. Young infants are protected from measles infection by maternal antibodies. Maternal Human Immunodeficiency Virus (HIV) infection may reduce levels of measles antibodies in newborns due to reduced placental transfer. Total IgG has been reported to significantly increase in HIV infected individuals due to polyclonal activation hence mutagenic activation. ART exposure in HIV infection causes a significant boost to immunity of the mother which is subsequently transferred to the infant. The study was a nested prospective cohort study within Kesho Bora Study that was conducted between 2005 – 2009 in which HIV infected pregnant women were enrolled and exposed to different anti-retroviral drugs depending on the degree of immune suppression. This study aimed at comparing total and anti-measles IgG levels and determining changes in the level of IgG in HIV infected women before and after ART. The correct dilution for Total IgG needed to be determined and the failing measles assays quality controlled before performing the tests. Total IgG assays involved testing using the recommended dilution factor of 1:8x104 which produced sample OD’s 2.9-3.6 this were high and could not be interpolated from best fit standard calibration curves. Dilution used in this study, was1:4x105aftermultiple dilutions andcalculation of standard error that produced tight error bars of +/-0.1 hence testing was done in singles. Cryopreserved maternal plasma was tested for TIgG and antimeaslesIgG using commercial ELISA kits before and 4 weeks after exposure to ARVs.The data was analyzed using SPSS software version 12.0. Paired t-test was used to compare the changes for IgG levels. The findings were that in 164 HIV pregnant women the mean of TIgG was 22.7g/l before and 17.11g/l after ART meaning there was a significant drop p-value was 0.000.7% of samples done for antimeaslesIgG tested equivocal which was significant compared to 5% negative andwas 2549.94 mIU/ml before and 2412.1mIU/ml after ART, p-value was 0.001hence there was a significant drop in the titres after ARVS administration. The findings will give insight on effects of exposure to ART on TIgG and antimeaslesIgG which will be useful for revision of current infant measles immunization schedules on children born to HIV seropositive mothers.