Fluorescent Antibody To Membrane Antigen Assay Research Articles

Measurement of antibodies to varicella-zoster virus using a virus-free fluorescent-antibody-to-membrane-antigen (FAMA) test.

The fluorescent-antibody-to-membrane-antigen (FAMA) test is regarded as the "gold standard" to detect protective antibodies to varicella-zoster virus (VZV) because of its high sensitivity and specificity. Because the classic FAMA test uses an infectious virus for detection of antibodies to VZV, it is labor-intensive, and also requires special equipment for handling the virus. For this reason, we attempted to develop a simple and safe FAMA assay. Because VZV glycoprotein E (gE) is one of the major VZV glycoproteins, we used the gE protein for the FAMA test (gE FAMA). Here, we demonstrate that overexpression of gE in HEK293T cells can be used to measure antibodies in human serum, and that gE FAMA titers are closely correlated with gpEIA ELISA data. These results indicate that our gE FAMA test has the potential to measure antibodies to VZV.

Evaluation Methods for the Immunogenicity of Varicella and Zoster Vaccines

Varicella vaccine has been included in the national immunization program for children since 2005 and zoster vaccine has been released since 2012 in Korea. Even though both varicella and zoster are caused by varicella-zoster virus (VZV), pathogeneses are different. In varicella, neutralizing antibody is very important to protect disease because VZV spreads via blood or lymph. In contrast, cell-mediated immunity is more important in zoster because of the neuronal spread of VZV. Therefore, the measurement methods of the immunogenicity against varicella and zoster vaccines are different. Fluorescent antibody to membrane antigen (FAMA) assay is the gold standard method to detect the protective antibody against VZV. It is still used as a reference test for the other methods. However, the fastidious nature required to perform the FAMA assay limits its use as a routine assay for the evaluation of vaccine immunogenicity. Nowadays, glycoprotein ELISA (gpEIA) is used as an alternative method for FAMA assay. However, there is no agreement over the protective level of gpEIA antibody titer with WHO standard international unit. The immunogenicity of zoster vaccine has been evaluated by responder cell frequency assay and IFN-γ ELISpot assay. Nevertheless, skin test is considered to be a more accurate biomarker for cell-mediated immunity against zoster. For the evaluation of varicella vaccine, it is necessary to standardize the FAMA assay and to set the cut-off value for the gpEIA antibody titer through long-term follow-up study. For zoster vaccine, the evaluation of cell-mediated immunity in Korean adults is urgently needed.

Comparative Study of the Standard Fluorescent Antibody to Membrane Antigen (FAMA) Assay and a Flow Cytometry-Adapted FAMA Assay To Assess Immunity to Varicella-Zoster Virus

A flow cytometry-adapted fluorescent antibody to membrane antigen (FAMA) assay to detect IgG antibodies against varicella-zoster virus (VZV) was developed and tested in 62 serum samples, showing 90.32% accuracy obtained from a receiver operating characteristic (ROC) curve with a 0.9125 (95% confidence interval [CI], 0.829 to 1.00) area below the curve compared to the result with standard FAMA.

Seroprevalence rate after one dose of varicella vaccine in infants

Live, attenuated varicella vaccine has been used since 1988 in Korea. However, varicella is still prevalent among both vaccinated and unvaccinated individuals, despite a relatively high level of immunization rate up to 80%. A recent report has demonstrated approximately 20% of primary vaccine failure rate after one dose of varicella vaccine using the fluorescent antibody to membrane antigen (FAMA) assay. The seroprevalence rate was determined using the FAMA and ELISA assays in 67 Korean infants following one dose of varicella vaccine. Positive fluorescence at a serum dilution of 1:4 or greater was considered as seropositive. The median age at vaccination was 12 months and the post-immunization sera were obtained on average 6.3 months (range 6 weeks-12 months) after vaccination. Among the 67 vaccinated infants, 56 (83.6%) were seropositive by FAMA assay while 30 (44.8%) were seropositive by ELISA. The geometric mean titers (GMTs) of the seropositive vaccinated infants (n = 56) were significantly lower than the GMTs of 9 individuals with a history of varicella (1:17.0 vs. 1:74.7, P = 0.001). Although there were no significant differences in seropositive rates according to intervals, there was a decreasing trend in the GMTs over time among the 56 seropositive recipients (r(2) = 0.154, P < 0.001). These data can be useful for optimizing the immunization strategy against varicella and should be confirmed by a prospective study including a large number of immunized infants.