Evaluation of the time resolved fluorescence immunoassay (TRFIA) for the detection of varicella zoster virus (VZV) antibodies following vaccination of healthcare workers

Abstract

Determination of varicella zoster virus (VZV) immunity in healthcare workers without a history of chickenpox is important for identifying those in need of vOka vaccination. Post immunisation, healthcare workers in the UK who work with high risk patients are tested for seroconversion. To assess the performance of the time-resolved fluorescence immunoassay (TRFIA) for the detection of antibody in vaccinated as well as unvaccinated individuals, a cut-off was first calculated. VZV-IgG specific avidity and titres six weeks after the first dose of vaccine were used to identify subjects with pre-existing immunity among a cohort of 110 healthcare workers. Those with high avidity (≥ 60%) were considered to have previous immunity to VZV and those with low or equivocal avidity (<60%) were considered naive. The former had antibody levels ≥ 400 mIU/mL and latter had levels < 400 mIU/mL. Comparison of the baseline values of the naive and immune groups allowed the estimation of a TRFIA cut-off value of > 130 mIU/mL which best discriminated between the two groups and this was confirmed by ROC analysis. Using this value, the sensitivity and specificity of TRFIA cut-off were 90% (95% CI 79-96), and 78% (95% CI 61-90) respectively in this population. A subset of samples tested by the gold standard Fluorescence Antibody to Membrane Antigen (FAMA) test showed 84% (54/64) agreement with TRFIA.

Fig. 1

Scatter plot to show the relationship between TRFIA titres and avidity (relative avidity index) RAI. (A) Six weeks post first vaccination; the dashed horizontal and vertical lines represent the avidity and TRFIA cut-offs (60% and log₁₀ 2.60;400 mIU/mLrespectively). Two results which did not cluster within the two populations are circled (1 and 2). (B) Antibody and avidity results six weeks following the second vaccination.

Fig. 2

Observed and fitted positive and negative distributions of baseline samples classified by avidity readings and TRFIA titres after the first dose of vaccine (six weeks). The arrow indicates the cut-off of log₁₀2.11 (130 mIU/mL); the point where the two fitted populations intercept. N=96. The horizontal dotted lines highlight the population that were TRFIA negative at baseline.

Fig. 3

ROC analysis of baseline TRFIA at different cut-offs. The area under the curve is 0.92.

Fig. 4

Relationship between FAMA scores and log₁₀ TRFIA readings. Whiskers extend to largest or smallest data point within 1.5 times the upper or lower quartile. The mean is indicated by a filled square (■) and the median by the horizontal line within the box. The horizontal dashed line indicates the log₁₀ TRFIA cut-off of 2.11 (130 mIU/mL). On analysis of variance with Tukey’s post hoc tests, the difference between the mean of the FAMA ≤2 and FAMA4groups was not significant (p>0.1): all other differences were significant at the 0.1% level.