For in vitro research use only, not for clinical diagnosis!
Bovine double-loaded protein (AMPH) ELISA kit.
Bovine double-loaded protein (AMPH) ELISA kit.
[Sample Handling and Requirements].
1. Serum: The whole blood specimen collected in the serum separation tube is placed at room temperature for 2 hours or 4 overnight, and then centrifuged at 1000 g for 20 minutes, and the supernatant can be taken or stored at -20 or -80, but repeated freezing and thawing should be avoided.
2. Plasma: Collect specimens with EDTA or heparin as anticoagulants, and centrifuge the specimens at 2-8 1000 g for 15 minutes within 30 minutes after collection, take the supernatant for detection, or store the supernatant at -20 or -80, but avoid repeated freezing and thawing.
3. Tissue homogenization: use pre-cooled PBS (001m, ph=7.4) Rinse the tissue, remove residual blood (lysed red blood cells in the homogenate will affect the measurement results), and mince the tissue after weighing. The minced tissue is compared to the corresponding volume of PBS (generally according to the weight to volume ratio of 1:9, for example, 1g of tissue sample corresponds to 9ml of PBS, and the specific volume can be appropriately adjusted according to the needs of the experiment, and a record should be made. It is recommended to add protease inhibitors to PBS) to a glass homogenizer and grind well on ice. To further lyse the histiocytes, the homogenate can be sonicated, or repeatedly freeze-thaw. After centrifugation of the homogenate at 5000 g for 5 10 minutes, the supernatant was taken for detection.
4. Cell culture supernatant or other biological specimens: centrifuge at 1000 g for 20 minutes, take the supernatant for detection, or store the supernatant at -20 or -80, but avoid repeated freezing and thawing.
Note: Hemolysis of the specimen will affect the results of the subsequent test, so the hemolyzed specimen should not be tested for this test.
Bovine double-loaded protein (AMPH) ELISA kit.
[Reagent Preparation].
The experimental procedure should be taken out of the refrigerated environment and should be used only after equilibration at room temperature.
20 Dilution of wash buffer: Distilled water is diluted 1:20, i.e., 1 part of 20 wash buffer plus 19 parts of distilled water.
Operation steps] 1, take out the required slats from the aluminum foil bag after 20min of room temperature equilibration, and seal the remaining slats with a ziplock bag and put them back 4.
2. Set up standard wells and sample wells, and add 50 L of standard with different concentrations to each standard well
3. Add 50 l of the sample to be tested to the sample wellBlank holes are not added.
4. Except for the blank wells, 100 L of horseradish peroxidase (HRP)-labeled detection antibody was added to each well of the standard and sample wells, and the reaction wells were sealed with a sealing film, and the reaction wells were incubated in a 37 water bath or incubator for 60min.
5. Discard the liquid, pat dry on the absorbent paper, fill each well with washing solution (350 l), let it stand for 1min, shake off the washing liquid, pat dry on the absorbent paper, and repeat the washing plate 5 times (you can also use a washing machine to wash the plate).
6. Add 50 L of substrate A and 50 L of substrate B to each well, and incubate for 37 min in the dark.
7. Add 50 L of stop solution to each well, and measure the OD value of each well at a wavelength of 450nm within 15min.
Bovine double-loaded protein (AMPH) ELISA kit.
[Procedure].
1.Dilution of standards: This kit provides a primary standard, which can be diluted in a small test tube according to the chart in the accompanying instructions.
2.Filling: Blank wells are set up respectively (blank control wells do not add samples and enzyme labeling reagents, and the rest of the steps are the same), standard wells, and sample wells to be tested. Accurately add 50 l of the standard on the microplate plate, add 40 l of sample diluent to the sample well, and then add 10 l of the sample to be tested (the final dilution of the sample is 5-fold). Add the sample to the bottom of the well of the microplate plate as much as possible, try not to touch the wall of the well, and gently shake to mix.
3.Incubation: Seal the plate with a sealing film and leave it for 37 minutes after incubation.
4.Preparation: Dilute the 30x concentrated washing solution 30 times with distilled water and set aside.
5.Washing: Carefully remove the sealing film, discard the liquid, spin dry, fill each well with washing liquid, let stand for 30 and then discard, repeat 5 times, pat dry.
6.Add enzymes: Add 50 l of enzyme labeling reagent per well, except for blank wells.
7.Incubation: The operation is the same as 3.
8.Washing: Same as 5.
9.Color development: Add chromogenic agent A50 L to each well, then add chromogenic agent B50 L, mix gently, and 37 avoid light for 10 minutes.
10.Termination: Add 50 l of stop solution per well to stop the reaction (blue turns yellow at this point).
11.Determination: The absorbance (OD value) of each well was measured sequentially with zero adjustment of the blank wells and the wavelength of 450 nm. The assay should be performed within 15 min of the addition of the stop solution.
Bovine double-loaded protein (AMPH) ELISA kit.
Bovine double-loaded protein (AMPH) ELISA kit.
[After-sales].
After-sales kits: The company's first-class kits are guaranteed in quality and quantity, and quality problems can be returned and exchanged free of charge, and free testing services are also provided.
Cell after-sales: 1. Cell transportation loss, bottle damage, serious leakage of culture medium, etc., reoccurre;
2. Please take photos on the day of receipt of the cell and on the 2nd and 3rd days, and those who are not informed will be deemed to be qualified. If there is a problem within 4-10 days, please provide the detailed steps of the cell** and the cell-related operation**, and communicate with my company's personnel in time to determine whether to reissue, you can refer to my official website or the relevant after-sales terms of the accompanying instructions.