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. 2014 Jul 2;20:956–969.

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Copyright © 2014 Molecular Vision.

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited, used for non-commercial purposes, and is not altered or transformed.

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First screening of CD4 clones for cytokine secretion upon stimulation with melanocytic proteins. Bacteria coated with complement were grouped in two pools, added to autologous Epstein-Barr virus-transformed B cells (EBV-B cells), and incubated overnight. CD4 T cells (5,000/well) were added in monoplicate to EBV-B cells (30,000/well). The amount of tumor necrosis factor (TNF)-α, interferon (IFN)-γ, interleukin (IL)-5, and IL-17 in the supernatant of overnight co-cultures was estimated with a multiplex bead-based assay. As the positive control, anti-MAGE-3 T cell clone R12C9 was used.