PRINCIPLE OF THE ASSAY

The Rheumera™ assay measures native type II collagen from cell/tissue culture or cartilage tissue using enzyme-linked immunosorbent assay (ELISA). The kit will measure 80 individual samples or 40 samples in duplicate. Removable strips and aliquoted reagents allow for samples to be tested on partial plates on 2 separate occasions. Samples are incubated in wells that are coated with antibodies to type II collagen. Dilutions of a standard (highly purified type II collagen) is also incubated in wells that are coated with antibodies to type II collagen. The wells are washed to remove unbound type II collagen molecules, and then incubated with antibodies to type II collagen which are conjugated to biotin. The wells are washed to remove unbound antibodies and then incubated with avidin (which has affinity to biotin) conjugated to peroxidase. The wells are washed to remove unbound avidin-peroxidase and then incubated with a chromogen substrate solution, TMB. A blue color develops which then turns yellow when the stop solution, mild sulfuric acid, is added. The color intensity is proportion to the amount of type II collagen present in the sample. The sample values, ng/ml of native type II collagen, are determined by the standard curve.

REAGENTS INCLUDED

1. Anti-type II collagen-coated strips (10).
2. Standard coated strips (4).
3. Diluent Buffer 1X (60ml). Solution
4. Wash Buffer 10X (50ml). Solution
5. Anti-type II collagen (2 vials). Lyophilized. Conjugated to biotin.
6. Standard (1 vial). Purified type II collagen 0.2 ml (1000 ng/ml).
7. Sample/Standard Dilution Buffer 1X (20ml). Solution
8. Avidin-Peroxidase (2 vials). Lyophilized
9. TMB (1X) - 10ml. Chromogen substrate for peroxidase detection.
10. Stop Solution (1X) – 5ml. Diluted sulfuric acid to stop the color reaction.

REAGENTS NOT INCLUDED

• Pepsin
• Elastase

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For Research Use Only, Not for Diagnostic or Therapeutic Use