Shanghai Xitang Biotechnology Co., Ltd. 021-55229872, 65333639
Rat kidney injury molecule (Kim-1) ELISA kit instruction manual
 ( for use in serum, plasma, cell culture supernatants, and urine biological fluids )
principle
This experiment used double antibody sandwich ABC-ELISA. The anti-rat Kim-1 monoclonal antibody was coated on the microtiter plate, Kim-1 of the standard and the sample was combined with the monoclonal antibody, and biotinylated anti-rat Kim-1 was added to form an immune complex attached to the plate. On the horseradish peroxidase-labeled Streptavidin combined with biotin, the substrate working solution is blue, and finally the stop solution sulfuric acid is added, and the OD value is measured at 450 nm. The Kim-1 concentration is proportional to the OD value, and can be passed. A standard curve was drawn to determine the Kim-1 concentration in the specimen.
Kit composition ( 2-8 ° C preservation)
Coated Wells | 96 holes | Enzyme Conjugate | 12ml |
10× specimen dilution (Sample Buffer) | 12ml | 20×Wash Buffer | 50ml |
Standards: 10ng/bottle | 2 bottles | Substrate working fluid (TMB Solution) | 12ml |
Primary antibody working solution (Biotinylated Antibody) | 12ml | Stop Solution | 12ml |
Prepare reagents and collect blood samples
1. Collect specimens: serum, plasma (heparin anticoagulation), cell culture supernatant, urine, etc., as soon as possible, store at 2-8 ° C for 48 hours; longer time must be frozen (-20 ° C or -70 ° C) to save, Avoid repeated freezing and thawing. Serum, plasma, as 1:20 dilution (10 ul of taking, sample plus diluent 190ul, diluted 20 times). The cell culture supernatant can be directly detected without dilution. The urine was diluted 2 times.
2. Standard solution preparation: Add 0.5 ml of distilled water before use and mix to prepare a 20 ng/ml solution. Set 8 tubes of standard tube, and add 200 ul of standard dilution solution to each tube. Add 20 ng of standard solution of 20 ng/ml to the first tube, mix and aspirate 200 ul with a pipette, and transfer to the second tube. Repeat the dilution as described above, and aspirate 200 ul from the seventh tube and discard it. The eighth tube is a blank control.
3. The 10× specimen dilution was diluted 1:10 with distilled water (example: 1 ml concentrated dilution + 9 ml distilled water).
4. Washing solution: diluted 1:20 with distilled water (example: 1 ml concentrated washing solution added to 19 ml of distilled water)
Test procedure
1. Loading: Add 100 ul of standard or sample to be tested in each well. Mix the reaction plate thoroughly and let it stand at 37 °C for 120 minutes.
2. Wash the plate: Wash the plate thoroughly with washing solution 4-6 times, and dry it on the filter paper.
3. Add 100 ul of the first antibody working solution to each well. The reaction plate was thoroughly mixed and placed at 37 ° C for 60 minutes.
4. Wash the board: the same as before.
5. Add 100 ul of enzyme-labeled antibody working solution per well. The reaction plate was placed at 37 ° C for 30 minutes.
6. Wash the board: same as before.
7. Add 100 ul of substrate working solution to each well and let it react at 37 ° C for 15 minutes in the dark.
8. Add 100 ul of stop solution to each well and mix.
9. Measure the absorbance at 450 nm using a microplate reader within 30 minutes.
Result calculation and judgment
1. All OD values ​​should be subtracted from the blank value before calculation.
2. Take the standard products 10000, 5000, 2500, 1250, 625, 312, 156, 0 pg/ml as the abscissa and OD as the ordinate. Draw on the coordinate paper and draw the standard curve.
3. Find the corresponding Kim-1 content on the graph based on the OD value of the sample, and multiply by the dilution factor.
Kit performance
1. Sensitivity: The minimum Kim-1 detection concentration is less than 80pg/ml.
2. Specificity: Recombinant or natural rat Kim-1 can be detected simultaneously. Does not cross-react with other cytokines in rats.
3. Repeatability: The coefficient of variation in both the plate and the plate is less than 10%.
Precautions
1. It is recommended to make double holes for the above standard holes and samples to be tested. The standard curve should be made at the same time for each measurement.
2. The washing process is critical. Insufficient washing will result in an accuracy error and an erroneous rise in the OD value.
3. After the slats are opened, the remaining slats should be sealed again to keep the slats dry .
4. This kit should be stored in a 4oC refrigerator.
5. This kit is for scientific research only and cannot be used for clinical diagnosis!
Hebei Abiding Co.Ltd, a professional food supplier, was established in 2005 in Shijiazhuang, China. We specialize ourselves in supplying Organic Tomato Paste and Organic Pasta. Conventional ones are also available to supply. We can make the private labels according to the customers' requirement. Our products have EU, NOP and JAS organic certificates. They are also Kosher certified. We have business relationships with importers and wholesalers from around the world, mainly in Europe and America. Over the years, we have earned highly recognition from clients due to our qualified service and steadily increase our market share year after year.
Products and Services
1. OEM service provided. We can do OEM with your brand(print, graphics on the products).
2. We have different types of organic pasta.(One-shop service)
3. International certificates. Guarantee our quality and make sure you can sell them all over the world!
Quinoa Pasta,Organic Gluten Free Quinoa Noodle,Bio Gluten Free Quinoa Noodle,Bio Gluten Free Quinoa Pasta
Hebei Abiding Co.Ltd , https://www.hebeiabiding.com