Effect of sample handling on analysis of cytokine responses to Mycobacterium tuberculosis in clinical samples using ELISA, ELISPOT and quantitative PCR.
Measuring cytokine responses to infection has proven to be invaluable for the understanding of immunity to tuberculosis in the laboratory. However, far less data are available from studies in humans and these have often produced conflicting results. Here we describe a comprehensive multi-center comparison of the most commonly used protocols for cytokine analysis: ELISA, ELISPOT and RT-PCR, in cohorts of TB patients, their household contacts and community controls. In particular, we have studied the effect on these protocols of conditions that commonly prevail in field studies, such as delays between sample collection and analysis, or different source material, such as whole blood or frozen PBMC. The results clearly show that while there is good correlation between the methods under optimal conditions, each method has strengths and weaknesses that render them more or less suitable for particular types of analyses. Researchers should carefully consider these factors when planning human field studies.
Doherty
T Mark
TM
Department of Tuberculosis Immunology, Statens Serum Institute, Copenhagen, Denmark. TMD@ssi.dk
Demissie
Abebech
A
Menzies
Dick
D
Andersen
Peter
P
Rook
Graham
G
Zumla
Alimuddin
A
D000293:Adolescent; D000328:Adult; D000818:Animals; D016207:Cytokines / Q000097:blood* / Q000276:immunology; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006801:Humans; D007118:Immunoassay; D008297:Male; D009169:Mycobacterium tuberculosis / Q000276:immunology*; D015203:Reproducibility of Results; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D013048:Specimen Handling
D016207:Cytokines