Poorly soluble peptides can mimic authentic ELISPOT responses.

R Karl Karlsson; Wim Jennes; Kimberly Page-Shafer; Douglas F Nixon; Barbara L Shacklett

doi:10.1016/j.jim.2003.11.013

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Poorly soluble peptides can mimic authentic ELISPOT responses.

R Karl Karlsson·Wim Jennes·Kimberly Page-Shafer·Douglas F Nixon·Barbara L Shacklett

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Abstract

The ELISPOT assay is a specific, sensitive, quantitative assay for assessing cell-mediated immune responses to a variety of antigens including HIV-1 peptides. In an interferon (IFN)-gamma-ELISPOT assay, peripheral blood mononuclear cells (PBMC) from two HIV-1 exposed seronegative (ESN) individuals appeared to respond strongly to an HIV Gag peptide. Analysis of this peptide revealed that it was incompletely dissolved and induced non-specific spot formation, even in the absence of cells. In subsequent experiments, the peptide was found to interact with avidin and the ELISPOT membrane. Filtering the peptide prevented non-specific spot formation. These findings underscore the need for appropriate controls and proper peptide preparation in order to reduce the risk of false-positive ELISPOT responses.

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Amino Acid Sequence Enzyme-Linked Immunosorbent Assay False Positive Reactions Gene Products, gag HIV Seronegativity HIV-1 Humans Interferon-gamma Peptides Solubility Journal Article Research Support, Non-U.S. Gov't Research Support, U.S. Gov't, P.H.S.

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R Karl Karlsson, Wim Jennes, Kimberly Page-Shafer, Douglas F Nixon, Barbara L Shacklett. 2004-02-01. Poorly soluble peptides can mimic authentic ELISPOT responses.. https://doi.org/10.1016/j.jim.2003.11.013

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