Investigation into Low-Level Anti-Rubella Virus IgG Results Reported by Commercial Immunoassays

Abstract

Since the 1980s, commercial anti-rubella virus IgG assays have been calibrated against a WHO International Standard and results have been reported in international units per milliliter (IU/ml). Laboratories testing routine patients' samples collected 100 samples that gave anti-rubella virus IgG results of 40 IU/ml or less from each of five different commercial immunoassays (CIA). The total of 500 quantitative results obtained from 100 samples from each CIA were compared with results obtained from an in-house enzyme immunoassay (IH-EIA) calibrated using the WHO standard. All 500 samples were screened using a hemagglutination inhibition assay (HAI). Any sample having an HAI titer of 1:8 or less was assigned a negative anti-rubella virus antibody status. If the HAI titer was greater than 1:8, the sample was tested in an immunoblot (IB) assay. If the IB result was negative, the sample was assigned a negative anti-rubella virus IgG status; otherwise, the sample was assigned a positive status. Concordance between the CIA qualitative results and the assigned negative status ranged from 50.0 to 93.8% and 74.5 to 97.8% for the assigned positive status. Using a receiver operating characteristic analysis with the cutoff set at 10 IU/ml, the estimated sensitivity and specificity ranged from 70.2 to 91.2% and 65.9 to 100%, respectively. There was poor correlation between the quantitative CIA results and those obtained by the IH-EIA, with the coefficient of determination (R 2 ) ranging from 0.002 to 0.413. Although CIAs have been calibrated with the same international standard for more than 2 decades, the level of standardization continues to be poor. It may be time for the scientific community to reevaluate the relevance of quantification of anti-rubella virus IgG.

Assessing Immunity to Rubella Virus: a Plea for Standardization of IgG (Immuno)assays

“…Under these conditions, RV IgG quantitative results and their interpretations may be different and even discordant for the same serum, depending on the CIA used. Discrepancies between assays have been confirmed by several studies (13)(14)(15) and can have an impact on diagnosis. Indeed, such discrepancies can lead to (i) confused clinical management of pregnant women, (ii) unnecessary revaccinations of already immune individuals, and (iii) reporting of false seroconversions among people with low RV IgG titers.…”

How to determine protective immunity in the post-vaccine era

“…As all of the currently available commercial screening assays detect total rubella IgG levels, and do not detect the level of neutralizing antibody (used to determine the recommended cutoff of 10 IU/ml), 12 the level of true protective antibody levels may not be accurately measured by commercial screening assays. Significant variability in precision and reproducibility across manufacture's platforms was observed when identical samples were tested on 5 different assays, 18 suggesting that a single cut off may not be able to be extrapolated to all rubella IgG screening assays. Additionally, while total IgG assays correlate well with high rubella IgG levels, the reproducibility of low rubella IgG titers, and particularly those around the cutoffs, is poor.…”

“…Additionally, while total IgG assays correlate well with high rubella IgG levels, the reproducibility of low rubella IgG titers, and particularly those around the cutoffs, is poor. 18 Large scale studies examining the level of neutralizing antibody in vaccinated populations who have achieved elimination are needed to understand the effect of waning total rubella IgG levels in the population. In light of changing epidemiology of universally vaccinated populations, and the variability between rubella IgG assays, it may therefore be appropriate to once again review the current indicators for immunity, and reexamine the role for neutralization assays in population screening.…”

Commentary: Where to Now for Standardization of Anti-Rubella Virus IgG Testing

“…The consequence of this poor standardization is that a patient's results are dependent upon what test kit is used. A person may obtain a negative result from one test kit and a positive result from another (4,5,10). A false-positive result may provide an incorrect sense of protection, whereas a false-negative result may cause an unnecessary vaccination.…”