Rat deoxypyridinol elisa kit instruction manual

Rat deoxypyridinoline Elisa kit instruction manual

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Rat elisa kit use purpose:
This kit is used to determine the content of deoxypyridinoline (DPD) in rat serum, urine and related liquid samples.
Rat elisa kit experimental principle This kit uses the double antibody sandwich method to determine the level of deoxypyridinoline (DPD) in the specimen. The microplate was coated with purified rat deoxypyridinoline (DPD) antibody to prepare a solid phase antibody, and deoxypyridinoline (DPD) was sequentially added to the microcapsule of the coated monoclonal antibody, followed by HRP-labeled deoxypyridinoline ( The DPD) antibody binds to form an antibody-antigen-enzyme-labeled antibody complex, which is thoroughly washed and then added to the substrate TMB for color development. TMB is converted to blue under the catalysis of HRP enzyme and converted to the final yellow color by the action of an acid. The color depth is positively correlated with deoxypyridinoline (DPD) in the sample. The absorbance (OD value) was measured with a microplate reader at a wavelength of 450 nm, and the concentration of rat deoxypyridinoline (DPD) in the sample was calculated from a standard curve.
Specimen requirements
1. The specimens should be extracted as soon as possible after collection, and the extraction should be carried out according to the relevant literature. The experiment should be carried out as soon as possible after extraction. If the test cannot be performed immediately, the specimen can be stored at -20 °C, but repeated freezing and thawing should be avoided.
2. Samples containing NaN3 could not be detected because NaN3 inhibited horseradish peroxidase (HRP) activity.
Operation Procedure Standard dilution: This kit provides one original standard, which can be diluted in a small tube according to the following chart.
Adding samples: blank holes (the blank control wells are not added with the sample and the enzyme standard reagent, the other steps are the same), the standard holes, and the sample holes to be tested. Accurately load 50 μl of the standard on the enzyme-labeled plate, add 40 μl of the sample dilution to the well to be tested, and then add 10 μl of the sample to be tested (the final dilution of the sample is 5 times). Add the sample to the bottom of the well of the microplate, try not to touch the wall of the well, and shake gently to mix.
Incubation: The plate was sealed with a sealing film and incubated at 37 ° C for 30 minutes.
Solution: 30 times concentrated washing solution is diluted with distilled water 30 times and then washed. Carefully remove the sealing film, discard the liquid, dry it, fill each well with washing liquid, let stand for 30 seconds, then discard it, repeat 5 Times, shoot dry.
Add enzyme: 50 μl of enzyme labeling reagent was added to each well, except for blank wells.
Incubation: The operation is the same as 3.
Washing: The operation is the same as 5.
Color development: Add 50 μl of color developer A to each well, then add 50 μl of color developer B, gently shake and mix, and develop color at 37 ° C for 15 minutes.
Termination: 50 μl of stop solution was added to each well to terminate the reaction (in this case, the blue color turned yellow).
Measurement: The absorbance (OD value) of each well was measured sequentially with a blank air conditioner of zero and a wavelength of 450 nm. The measurement should be carried out within 15 minutes after the addition of the stop solution.

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