Serum-free culture medium and IL-7 costimulation increase the sensitivity of ELISpot detection.

Archive ouverte

Martinuzzi, Emanuela | Scotto, Matthieu | Enée, Emmanuelle | Brezar, Vedran | Ribeil, Jean-Antoine | van Endert, Peter | Mallone, Roberto

Edité par CCSD ; Elsevier -

International audience. The identification of parameters maximizing detection sensitivity in ELISpot assays is important to transfer this technology into the clinical setting for identifying rare Ag-specific CD8(+) T cells. We have therefore considered human IFN-gamma CD8(+) T cell responses against viral epitopes to analyze different variables which could be critical during the epitope-specific stimulation period. Two parameters were found to greatly enhance detection sensitivity (i.e., to specifically increase epitope-driven signal while keeping background noise to a minimum): use of human serum-free vs. serum-supplemented culture medium (2.4-fold median increase) and addition of low dose IL-7 (1.5-fold increase). Incorporating both of these parameters into the ELISpot procedure proved capable of greatly amplifying (35.1-fold increase) the low grade CD8(+) T cell responses directed against beta-cell epitopes of type 1 diabetes patients, as compared to a previously optimized procedure using human serum-supplemented medium and low dose IL-2. Implementation of this ELISpot procedure should expedite development of "immune staging" protocols for autoimmune as well as tumor and infectious diseases.

• Description
• Sujet/Public
• Outil
• Outil d'anticipation
• Mémoire et thèse
• Gestion