Kit Instruction Manual This kit is for research use only. Kit composition The calculation takes the concentration of the standard as the abscissa and the OD value as the ordinate, and draws a standard curve on the coordinate paper. Linear range: Medical Cart,Med Cart,Mobile Medical Carts,Medical Trolley Cart Hebei Boxin Rehabilitation Equipment Co., Ltd , https://www.cnsmartcare.com
Drug Name:
Generic name: Human C-reactive protein (CRP) enzyme-linked immunoassay kit Purpose of use:
This kit is used to determine the content of C-reactive protein (CRP) in human serum, plasma and related liquid samples.
Experimental principle The kit uses an indirect method to determine the level of human C-reactive protein (CRP) in the specimen. Using purified human C-reactive protein (CRP)
The antibody is coated on the microwell plate to make a solid-phase antibody, and the known concentration of C-reactive protein (CRP) is sequentially added to the monoclonal antibody coated microwell
Standard and unknown concentration of C-reactive protein (CRP) to be tested, after incubation, add biotin-labeled anti-IgG antibody,
It is combined with streptavidin-HRP to form an immune complex. After thorough washing, the substrate TMB is added for color development. TMB at
The HRP enzyme catalyzes the conversion to blue and the acid to the final yellow. The shade of the color and the
C-reactive protein (CRP) was positively correlated. The absorbance (OD value) was measured with a microplate reader at a wavelength of 450 nm, and the concentration of human C-reactive protein (CRP) in the sample was calculated by a standard curve.
1 20 times concentrated washing solution 50ml × 1 bottle of standard product S1 (1800μg / L) 0.5ml × 1 bottle
2 Streptavidin-HRP 6ml × 1 bottle of standard product S2 (1200μg / L) 0.5ml × 1 bottle
3 Enzyme label coating plate 12 well × 8 standard S3 (600μg / L) 0.5ml × 1 bottle
4 Biotin-labeled anti-IgG antibody 6ml × 1 bottle of standard S4 (300μg / L) 0.5ml × 1 bottle
5 Developer A solution 6ml × 1 bottle
9
Standard product S5 (150μg / L) 0.5ml × 1 bottle
6 Developer B solution 6ml × 1 / bottle 10 sealed bag 1
7 Stop solution 6ml × 1 bottle 11 sealing film 3 sheets
8 Sample diluent 6ml × 1 bottle 12 Instructions 1 specimen requirement
1. The samples containing NaN3 cannot be detected because NaN3 inhibits horseradish peroxidase (HRP) activity.
2. Specimens are extracted as soon as possible after collection, and extraction is performed according to relevant literature, and experiments should be conducted as soon as possible after extraction. If the test cannot be performed immediately, the specimen can be stored at -20 ℃, but repeated freezing and thawing procedures should be avoided
1. Determine the number of slats required based on the number of samples to be tested plus the number of standard products. It is recommended to make multiple holes for each standard and blank hole. Each sample is determined according to its own quantity, and those that can use multiple holes can be used as much as possible.
2. Adding samples: set up blank wells (the blank control wells do not add samples, the rest of the operations are the same), standard wells, and sample wells to be tested. Then add 50μl of the standard to the standard well, add 10μl of the sample to be tested to the sample reaction well and then add 40μl of the sample diluent (the final dilution of the sample is 5 times), cover with the sealing film, and shake gently to mix, 37 ℃ Incubation
45 minutes.
3. Mixing solution: dilute 20 times concentrated washing solution with distilled water 20 times and reserve
4. Washing: Carefully peel off the sealing film, discard the liquid, spin dry, fill each well with the washing liquid, let it stand for 30 seconds and then discard, repeat this 4 times, pat dry. 5. Add biotin-labeled anti-IgG antibody: Add 50 μl of biotin-labeled anti-IgG antibody to each well. Incubate at 37 ° C for 30 minutes
6. Washing: The operation is the same as 4.
7. Add streptavidin-HRP: add 50μl streptavidin-HRP to each well, gently shake and mix, incubate at 37 ℃ for 30
minute.
8. Washing: The operation is the same as 4.
9. Color development: add 50μl of developer A to each well, and then add 50μl of developer B, mix gently, and avoid color development at 37 ℃
15 minutes.
10. Termination: Add 50μl of stop solution to each well to stop the reaction (at this time the blue will turn to yellow).
11. Determination: Measure the absorbance (OD value) of each well in sequence with the blank air conditioner at zero and 450nm wavelength. The measurement should be carried out within 15 minutes after adding the stop solution.
The corresponding concentration of OD value is found by the standard curve; then multiplied by the dilution factor; or the linear regression equation of the standard curve is calculated from the concentration of the standard and the OD value, and the OD value of the sample is entered into the equation to calculate the sample concentration, and then Multiplied by the dilution factor,
This is the actual concentration of the sample.
Precautions
1. The kit should be taken out of the refrigerated environment and equilibrated at room temperature for 15-30 minutes before use. If the enzyme-coated plate is unsealed after opening, the strip should be stored in a sealed bag.
2. Crystals may be precipitated in the concentrated washing liquid, which can be heated and dissolved in a water bath during dilution, and the results will not be affected during washing.
3. The sampler should be used at each step of sample addition, and the accuracy should be regularly checked to avoid test errors. It is best to control the sampling time within 5 minutes. If there are many specimens, it is recommended to use a volley gun to add samples.
4. If the content of the substance to be tested in the specimen is too high (the OD value of the sample is greater than the OD value of the first well of the standard well), please first dilute the sample by a certain multiple (n times) and then determine it. When calculating, please multiply the dilution factor (× 5 × n).
5. The sealing film is limited to one-time use to avoid cross-contamination.
6. The components of different batches of this reagent shall not be mixed. Store developer B in dark place.
7. Strictly follow the instructions, and the test results must be determined by the microplate reader.
8. All samples, washing liquids and various wastes should be treated as infectious agents.
9. If there is any difference with the English manual, the English manual shall prevail.
100μg / L -2000μg / L
specification:
96 servings / box storage conditions and expiration date
1. Kit storage :; 2-8 ℃.
2. Validity: 6 months
Human C-reactive protein (CRP) enzyme-linked immunoassay (ELISA)