Insulin - ELISA
PREPARATION FOR ASSAY
-Bring all reagents and samples to room temperature (20-25°C) and mix gently before beginning the test.
-Have all reagents and samples ready before the start of the assay. Once the test has begun, it must be performed without any interruption to get the most reliable and consistent results.
-Use new disposable tips for each specimen.
-Secure the desired number of coated wells in the holder. Mark date sheet with sample identification.
-Dispense 25 ul of serum sample, controls and reference into the assigned wells.
-Dispense 100 ul of Enzyme conjugate into each well and mix for 5 seconds.
-Incubate for 30 minutes at 25°C
-Remove incubation mixture and rinse the wells five times with washing buffer.
-Dispense 100 ul of Solution A and then 100 ul of Solution B into each well.
-Incubate for 15 minutes at R.T.
-Stop reaction by adding 50 ul of 1N sulfuric acid or 2N HCl to each well and read O.D. at 450 nm with a microwell reader.
1. Clark PMS & Hales CN (1991) Assay of Insulin. In P.C. Pickup and G.Williams eds. Textbook of Diabetes, Vol 1, 335-347, Blackwell Scientific Publications
2. Clark PMS and Hales CN (1994)How to Measure Plasma Insulin. Diabetes Reviews,10:79-90
3. Andersen L, Dinesen B. Jorgensen PN, Poulsen F and Roder MF (1993) Enzyme Immunoassay for Intact Human Insulin in Serum or Plasma. Clin Chem 38:578-582
4. Volund A (1993) Conversion of Insulin units to SI units. American Journal of Clinical Nutrition