Wednesday, October 29, 2003

Description
A simple ELISA assay to measure cytokines in culture supernatants

Procedure
- Coat plates with primary Antibody 2 g/ml, 50 l/well
- Incubate overnight 4C
- Wash 3 times
- Block with 10% FCS, 200 ml/well
- Incubate 2 hours room Temp.
- Add standards and sample, 100 ml/well
- Incubate 45 minutes room Temp.
- Wash 3 times
- Add secondary antibody (biotinylated) 1 g/ml, 100 l/well
- Incubate 45 minutes room Temp.
- Wash 3 times
- Add Avidin-peroxydase 1 g/ml, 100 l/well
- Incubate 30 minutes room Temp.
- Wash 3 times
- Add substrate ABTS (remember to thaw it and add 10 ml of H2O2 per 10 ml of substrate).
- Read 405 nm


Recipes


Supplies
Ab were purchased from R & D sytems
ELISA plates were Dynatech

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