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Immunological assays Tumor formation Tumor microenvironmentVaccine-induced Cytokine Production Detected by Luminex Multiplex Analysis
Different vaccine and adjuvant combinations are known to rapidly induce antigen presenting cell (APC) maturation and pro-inflammatory cytokine and production, which in turn play an important role in the priming of antigen-specific T cells. Measuring cytokine production systemically in the serum fails to detect localized responses in the lymph nodes draining a subcutaneous immunization site. On the other hand, stimulating APC with vaccine formulations in vitro lacks the complexity of the lymph node microenvironment and the presence of other in vivo factors. Here we analyse cytokine production directly in vaccine draining lymph nodes (dLN) extracted early after in vivo vaccination. To do this we perform cytokine multiplex analysis of supernatants from whole dLN cell suspensions following a brief ex vivo incubation.
Isolation of Mouse Tumor-Infiltrating Leukocytes by Percoll Gradient Centrifugation
A hallmark of cancer-associated inflammation is the infiltration of leukocytes into tumors, which is believed to be recruited by chemokines. Some infiltrating leukocytes such as macrophages often promote tumor growth by producing growth-inducing and angiogenic factors. Here, we described a method of isolating tumor-infiltrating leukocytes with Percoll density gradient, because Percoll possesses a low viscosity, a low osmolarity and no toxicity to cells. Different leukocyte populations are isolated based on their density and collected at the interface between 40% and 80% discontinuous Percoll gradient.