Cells derived from the nasal mucosa, spleen, or colon were stimulated for 4–5 hours with PMA (50 ng/ml), lonomycin (500 μg/ml) (Sigma Aldrich), and golgi stop (BD) (1 μg/ml), or stimulated overnight with WCA (10 μg/ml) followed by Golgistop (1 μg/ml) for 4–5 hours. Following these incubations, cells were harvested and prepared for surface and intracellular cytokine staining as described above. Nasal cell suspensions were aliquoted into 24-well TC plates in complete tissue culture media (TCM) and stimulated with WCA (10 μg/ml ) for 7 days. Supernatants were harvested and IL-17A was detected in the supernatants by ELISA as per the mouse IL-17A ELISA DUO set kit instructions (reference DY421; R&D Systems). IL-17A was detected in samples of WCA-stimulated whole blood as previously described18 . A value of 8 pg/ml was set as the lower limit of detection of IL-17A cytokine protein by ELISA and is represented by the dotted line in IL-17A cytokine detection graphs.
Mouse il 17a elisa duo set kit
Manufactured by R&D Systems
The Mouse IL-17A ELISA DUO set kit is a laboratory product designed to detect and quantify the concentration of mouse interleukin-17A (IL-17A) in various sample types. It is a sandwich enzyme-linked immunosorbent assay (ELISA) that uses a pair of specific antibodies to capture and detect the target analyte.
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2 protocols using mouse il 17a elisa duo set kit
1
Measuring IL-17A in Immune Cell Assays
2
Measurement of IL-17A Production
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Cells derived from the nasal mucosa, spleen, or colon were stimulated for 4–5 h with PMA (50 ng/ml), Ionomycin (500 ng/ml) (Sigma-Aldrich), and Golgistop (BD) (1 µg/ml), or stimulated overnight with WCA (10 μg/ml) followed by Golgistop (1 µg/ml) for 4–5 h. Following these incubations, cells were harvested and prepared for surface and intracellular cytokine staining as described above. Nasal cell suspensions were aliquoted into 24-well TC plates in complete tissue culture media (TCM) and stimulated with WCA (10 μg/ml) for 7 days. Supernatants were harvested and IL-17A was detected in the supernatants by ELISA as per the mouse IL-17A ELISA DUO set kit instructions (reference DY421; R&D Systems). IL-17A was detected in samples of WCA-stimulated whole blood as previously described.19 (link) A value of 8 pg/ml was set as the lower limit of detection of IL-17A cytokine protein by ELISA and is represented by the dotted line in IL-17A cytokine detection graphs.
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