Performance and Stability of New Class of Fetal Bovine Sera (FBS) and Its Lyophilized Form in ELISpot and FluoroSpot Assays: Applications for Monitoring the Immune Response in Vaccine, and Cell and Gene Immunotherapy in Clinical Trials.

Autor:	Hosseini Z; Translational Science and Innovation Laboratory (TSAIL), Q Solutions, Durham, NC, USA., Groves CJ; Translational Science and Innovation Laboratory (TSAIL), Q Solutions, Durham, NC, USA., Anders P; Translational Science and Innovation Laboratory (TSAIL), Q Solutions, Durham, NC, USA., Cave K; Translational Science and Innovation Laboratory (TSAIL), Q Solutions, Durham, NC, USA., Krunkosky M; Translational Science and Innovation Laboratory (TSAIL), Q Solutions, Durham, NC, USA., Chappell B; Translational Science and Innovation Laboratory (TSAIL), Q Solutions, Durham, NC, USA., Pattyn S; ImmunXperts, a Q Solutions Company, Gosselies, Belgium., Davis D; Seraworx, Providence, UT, USA., Janetzki S; ZellNet Consulting, Inc., Fort Lee, NJ, USA., Reap E; Translational Science and Innovation Laboratory (TSAIL), Q Solutions, Durham, NC, USA. Elizabeth.Reap@q2labsolutions.com.
Jazyk:	angličtina
Zdroj:	Methods in molecular biology (Clifton, N.J.) [Methods Mol Biol] 2024; Vol. 2768, pp. 305-316.
DOI:	10.1007/978-1-0716-3690-9_18
Abstrakt:	Interferon-gamma (IFNγ) ELISpot and FluoroSpot are widely used assays to detect functional cell responses in immunotherapy clinical studies. Recognized for their importance in vaccine development studies to quantitate immune responses, these assays have more recently risen to the forefront in cell and gene therapy as well as cancer immunotherapy fields where responses against cancer neoantigens are not easily detectable above assay background. Here, we test a new class of fetal bovine serum (FBS), CultraPure FBS, in ex vivo ELISpot and FluoroSpot assays and cultured FluoroSpot assays following in vitro expansion. Several CultraPure FBS lots that have been specially formulated through the process of lyophilization (lyo-FBS) were compared to liquid CultraPure FBS. We stimulated human PBMCs with antigen-specific peptide pools diluted in media supplemented with liquid CultraPure FBS or lyo-FBS and found equivalent cytokine production with negligible to no assay background with both liquid and lyo-FBS formats. Moreover, the lyo-FBS showed lot-to-lot consistency and 90-day refrigerated (4 °C) stability in both ex vivo direct and in vitro cultured assays. In addition, we present here a method using lyo-FBS for the expansion of low-frequency antigen-specific T cells, mimicking the low frequency seen with cancer neoantigens by utilizing a cultured FluoroSpot assay. Our results demonstrate the presence of Granzyme B, interferon-gamma (IFNγ), and tumor necrosis factor (TNF) production by antigen-specific polyfunctional T cells following a 9-day culture using media supplemented with lyo-FBS. (© 2024. The Author(s), under exclusive license to Springer Science+Business Media, LLC, part of Springer Nature.)
Databáze:	MEDLINE
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