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Methods Mol Biol. 2012;792:97-104. doi: 10.1007/978-1-61779-325-7_8.

ELISPOT assay for neuroscience research: studying TNFα secretion from microglial cells.

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R&D Systems, Inc., Minneapolis, MN, USA.


The major application of ELISPOT assays is to study secretion of cytokines and chemokines from immune system cells. We adapted this assay to study TNFα secretion from microglial BV2 cells, which are similar in physiology to microglia in the nervous system. Stimulation of BV2 cells with 1 μg/mL LPS resulted in a robust secretion of TNFα. Unlike uniform round spots formed by TNFα secreted by immune system cells, BV2 cells produced spots with short zigzag "tails" indicating that BV2 cells were actively moving during the incubation. In spite of irregular shapes, spots could be easily counted using an ELISPOT reader. Our study has shown the feasibility of employing an ELISPOT assay as a tool for neuroscience research to study the mechanisms underlying protein secretion from microglial cells. In addition, due to its convenient format, ELISPOT can be used for high-throughput screening of the potency of novel drugs to stimulate or inhibit cytokine secretion by microglial cells in the brain.

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