Rat C-Reactive Protein/CRP DuoSet ELISA Summary

Product Summary

This DuoSet ELISA Development kit contains the basic components required for the development of sandwich ELISAs to measure natural and recombinant rat C-Reactive Protein/CRP. The suggested diluent is suitable for the analysis of most cell culture supernate samples. Diluents for complex matrices, such as serum and plasma, should be evaluated prior to use in this DuoSet.

Preparation and Storage

Background: C-Reactive Protein/CRP

C-Reactive Protein (CRP), also known as Pentraxin 1, is a secreted pentameric protein that functions as a sensor and activator for the innate immune response. In humans, it is a major acute-phase protein; its circulating concentration is dramatically elevated at the onset of inflammation. In mice, however, serum CRP levels increase only slightly during inflammation, and the analogous acute phase role is filled by Pentraxin 2. CRP binds, opsonizes, and induces the phagocytosis of bacteria and apoptotic cells. It regulates activation of the classical complement pathway by binding several proteins in the complement cascade as well as Fc gamma RI, Fc gamma RIIA, and Fc gamma RIIB on macrophages and dendritic cells. It also promotes dendritic cell maturation and humoral immunity. In cardiovascular disease, CRP binds to oxidized LDL, exacerbates tissue damage in myocardial infarction, and inhibits the repair of injured vascular endothelium.

Assay Procedure

Refer to the product datasheet for the complete assay procedure.

Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.

1.     Prepare all reagents, standard dilutions, and samples as directed in the product insert.

2.     Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.

3.     Add 50 μL of Assay Diluent to each well.

4.     Add 50 μL of Standard, control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours.

5.     Aspirate each well and wash, repeating the process 4 times for a total of 5 washes.

6.     Add 100 μL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours.

7.     Aspirate and wash 5 times.

8.     Add 100 μL Substrate Solution to each well.

9.     Add 100 μL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.