A protocol for high-throughput screening for immunomodulatory compounds using human primary cells.

High-throughput screening is a powerful platform that can rapidly provide valuable cytotoxic, immunological, and phenotypical information for thousands of compounds. Human peripheral blood mononuclear cells (PBMCs) cultured in autologous plasma can model the human immune response. Here, we describe a protocol to stimulate PBMCs for 72 h and measure cytokine secretion via AlphaLISA assays and cell surface activation marker expression via flow cytometry. Cryopreserved PBMCs are incubated for 72 h with various small molecule libraries and the supernatants are harvested to rapidly measure secretion levels of key cytokines (tumor necrosis factor alpha, interferon gamma, interleukin 10) via the AlphaLISA assay. Almost simultaneously, the cells can be fixated and stained using antibodies against innate immune activation markers (CD80, CD86, HLA-DR, OX40) for analysis via flow cytometry. This multiplexed readout workflow can directly aid in the phenotypic identification and discovery of novel immunomodulators and potential vaccine adjuvant candidates. For complete details on the use and execution of this protocol, please refer to Chew et al.

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Copyright © 2023 The Author(s). Published by Elsevier Inc. All rights reserved.

Déclaration de conflit d'intérêts

Declaration of interests S.V.H., O.L., and D.J.D. are named inventors on vaccine adjuvant patents assigned to Boston Children’s Hospital. O.L. served as a paid consultant to Moody’s analytics. D.J.D. is on the scientific advisory board of EdJen BioTech and serves as a consultant with Merck Research Laboratories/Merck Sharp & Dohme Corp. (a subsidiary of Merck & Co., Inc.). These commercial or financial relationships are unrelated to the current study.