We have evaluated a novel B-cell FluoroSpot assay for the analysis of antibody responses in healthy individuals vaccinated intramuscularly with Influenza A (H1N1) antigen (Pandemrix®, GlaxoSmithKline). Using the FluoroSpot assay and an ELISpot assay run in parallel for comparison, we measured the frequency of cells secreting antigen-specific as well as total IgG or IgA antibodies seven days pos...

Diagnosis of tuberculous pleurisy remains a challenge in the clinic. In this study, we evaluated the usefulness of a previously developed Mycobacterium tuberculosis antigen-specific gamma interferon enzyme-linked immunospot (ELISPOT) assay in the diagnosis of tuberculous pleurisy by testing a cohort of 352 patients with pleural effusion. We found that M. tuberculosis antigen-specific gamma inte...

Cerebrospinal fluid (CSF) and peripheral blood (PBL) were sampled multiple times from 25 patients with a clinical diagnosis of tuberculous meningitis (TBM) and 49 controls, including 27 patients with other infectious diseases of the central nervous system and 22 patients with other noninfectious neurological diseases. We used an enzyme-linked immunospot assay (ELISPOT) to detect anti-Mycobacter...

Immune monitoring of functional responses is a fundamental parameter to establish correlates of protection in clinical trials evaluating vaccines and therapies to boost antigen-specific responses. The IFNg ELISPOT assay is a well-standardized and validated method for the determination of functional IFNg-producing T-cells in peripheral blood mononuclear cells (PBMC); however, its performance gre...

CD8 T-cell responses were examined in subjects with incident (new following negative visits) or prevalent (lasting ≥ 4 months) human papillomavirus type 16 (HPV16) or human papillomavirus (HPV18) infection. The groups were chosen from a cohort of women being followed every 4 months with cervical cytology and HPV-DNA testing. Enzyme-linked immunospot (ELISPOT) assay was performed at enrollment (...

Human immunodeficiency virus (HIV)'s tremendous sequence variability is a major obstacle for the development of cytotoxic-T-lymphocyte-based vaccines, especially since much of this variability is selected for by CD8(+) T cells. We investigated to what extent reactivity to escape variant peptides in standard enzyme-linked immunospot (ELISPOT) assays predicts the recognition of cells infected wit...

The plaque reduction neutralization test (PRNT) is used widely to measure the neutralization activity of anti-dengue virus (DENV) antibodies, but it is time-consuming and labor-intensive and has low sample throughput. For fast and convenient measurement of neutralizing antibodies, especially in evaluating the efficiency of the DENV vaccines on a large scale, a new method is needed to replace PR...

A number of different interferon-gamma ELISpot protocols are in use in laboratories studying antigen-specific immune responses. It is therefore unclear how results from different assays compare, and what factors most significantly influence assay outcome. One such difference is that some laboratories use a short in vitro stimulation period of cells before they are transferred to the ELISpot pla...

Tuberculosis is a global health problem. The Mycobacterium bovis Bacille Calmette Guerin (BCG) vaccine has variable efficacy (0-80%) so there is a drive to develop novel vaccines. The cytokine, interferon gamma (IFNγ), is an essential component of the protective response to M. tuberculosis (M. tb) infection and is also produced in response to BCG vaccination. Induction of an IFNγ response is us...

BACKGROUND The Comprehensive T Cell Vaccine Immune Monitoring Consortium (CTC-VIMC) was created to provide standardized immunogenicity monitoring services for HIV vaccine trials. The ex vivo interferon-gamma (IFN-γ) ELISpot is used extensively as a primary immunogenicity assay to assess T cell-based vaccine candidates in trials for infectious diseases and cancer. Two independent, GCLP-accredite...