Detection of keyhole limpet hemocyanin (KLH)-specific immune responses by intracellular cytokine assay in patients vaccinated with idiotype-KLH vaccine.
Keyhole limpet hemocyanin (KLH) has been used as an immune potentiator to enhance antigen-specific responses against haptens and weak antigens including self-antigens. In the present study, we describe the optimization of the intracellular cytokine response to KLH in peripheral blood mononuclear cells (PBMC) of lymphoma and myeloma patients that were vaccinated with tumor-specific immunoglobulin (Id) conjugated to KLH. Addition of anti-CD28 antibody significantly enhanced cytokine-producing CD4-T cells. While fresh PBMC showed maximal cytokine response 14 h after antigen stimulation, the frozen PBMC showed maximal cytokine responses by 24 h. Supplementation of the culture medium with fetal bovine serum gave a better signal-to-noise ratio than human AB serum in the intracellular detection of cytokines. The intracellular cytokine responses correlated with the cytokine measurements by enzyme-linked immunosorbent assay (ELISA). Together these results indicate that the intracellular cytokine assay is very helpful to measure antigen-specific immune responses, and in subsequent studies, we have utilized this sensitive technique to detect immune responses against tumor antigens such as idiotype.
Neelapu
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SS
ETI Branch, NCI at Frederick, Md 21702-1201, USA.
Baskar
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Kwak
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LW
D000951:Antigens, Neoplasm / Q000276:immunology*; D018106:CD28 Antigens / Q000276:immunology; D019496:Cancer Vaccines / Q000276:immunology*; D016207:Cytokines / Q000096:biosynthesis*; D004797:Enzyme-Linked Immunosorbent Assay; D006433:Hemocyanins / Q000276:immunology*; D006801:Humans; D007130:Immunoglobulin Idiotypes / Q000276:immunology*; D014611:Vaccination; D018074:Vaccines, Conjugate / Q000276:immunology
D000951:Antigens, Neoplasm; D018106:CD28 Antigens; D019496:Cancer Vaccines; D016207:Cytokines; D007130:Immunoglobulin Idiotypes; D018074:Vaccines, Conjugate; D006433:Hemocyanins; C032808:keyhole-limpet hemocyanin