The Adrenocorticotropic Hormone (ACTH) BioAssay™ ELISA Kit (Mouse) is a competitive inhibition immunoassay for the in vitro quantitative measurement of ACTH in mouse plasma, serum and other biological fluids.
Test Principle:This assay employs the competitive inhibition enzyme immunoassay technique. An antibody specific for ACTH has been pre-coated onto a microplate. A competitive inhibition reaction occurs between biotin-labeled ACTH and unlabeled ACTH in standards or samples for limited binding sites on the pre-coated antibody. After incubation, unbound materials are washed off. Next, avidin conjugated to horseradish peroxidase (HRP) is added to each microplate well and incubated. The amount of bound HRP conjugate is inversely proportional to the concentration of ACTH in the sample. After addition of the substrate solution, the intensity of color developed is inversely proportional to the concentration of ACTH in the sample.
Kit Components:*023172A: Microtiter Plate, 96 wells, Pre-coated, ready to use
Storage and Stability:Store *023172A, *023172B, *023172D and *023172E at -20°C. Store all the other components at 4°C. Unused kit is stable for 6 months. Once kit components are opened, it is highly recommended to use remaining reagents within 1 month provided this is within the expiration date of the kit. For maximum recovery of product, centrifuge the original vials after thawing and prior to removing the cap.
Assay Procedure Summary:1. Prepare all reagents, samples and standards
2. Add 50ul standard or sample to each well, then add 50ul Detection Reagent A immediately. Shake and mix. Incubate 1 hour at 37°C
3. Aspirate and wash 3 times
4. Add 100ul prepared Detection Reagent B. Incubate 30 minutes at 37°C
5. Aspirate and wash 5 times
6. Add 90ul Substrate Solution. Incubate 10-20 minutes at 37°C
7. Add 50ul Stop Solution. Read absorbance at 450nm immediately.
