Human prostaglandin F2α (PGF2α) enzyme-linked immunoassay (ELISA) kit instruction manual Xiamen Huijia Biotechnology Co., Ltd. This reagent is for research use only: this kit is used to determine prostaglandins in human serum, plasma and related liquid samples F2α (PGF2α) content. Experimental principle: This kit uses the double antibody sandwich method to determine the level of human prostaglandin F2α (PGF2α) in the specimen. The microplate was coated with purified human prostaglandin F2α (PGF2α) antibody to make a solid phase antibody. Prostaglandin F2α (PGF2α) was added to the monoclonal antibody-coated microwells in turn, followed by HRP-labeled prostaglandin F2α (PGF2α ) The antibody binds to form an antibody-antigen-enzyme-labeled antibody complex. After thorough washing, the substrate TMB is added for color development. TMB is converted into blue under the catalysis of HRP enzyme, and into the final yellow under the action of acid. The color depth is positively correlated with prostaglandin F2α (PGF2α) in the sample. The absorbance (OD value) was measured with a microplate reader at a wavelength of 450 nm, and the concentration of human prostaglandin F2α (PGF2α) in the sample was calculated by a standard curve. Kit composition: kit composition 48 well configuration 96 well configuration storage instructions 1 copy 1 copy sealing film 2 pieces (48) 2 pieces (96) 1 sealed bag 1 enzyme coated plate 1 × 48 1 × 96 2 Store standard at -8 ℃: 1350pg / ml 0.5ml × 1 bottle 0.5ml × 1 bottle 2-8 ℃ store standard dilution 1.5ml × 1 bottle 1.5ml × 1 bottle 2-8 ℃ store enzyme label reagent 3 ml × 1 bottle of 6 ml × 1 bottle of sample diluent at 2-8 ° C 3 ml × 1 bottle of 6 ml × 1 bottle of 2-8 ° C Developer A 3 ml × 1 bottle of 6 ml × 1 bottle of 2-8 ° C Developer B solution 3 ml × 1 bottle 6 ml × 1 bottle at 2-8 ° C Storage stop solution 3ml × 1 bottle 6ml × 1 bottle 2-8 ° C Store 20 times concentrated washing solution 20ml × 1 bottle 30ml × 1 bottle 2- Sample handling and requirements for storage at 8 ° C: 1. Serum: room temperature blood coagulates naturally for 10-20 minutes, centrifuged for about 20 minutes (2000-3000 rpm). Collect the supernatant carefully and centrifuge again if a precipitate appears during storage. 2. Plasma: EDTA, sodium citrate or heparin should be selected as the anticoagulant according to the requirements of the specimen, mixed for 10-20 minutes, and centrifuged for about 20 minutes (2000-3000 rpm). Collect the supernatant carefully. If a precipitate forms during storage, it should be centrifuged again. 3. Urine: collected in a sterile tube and centrifuged for about 20 minutes (2000-3000 rpm). Collect the supernatant carefully. If a precipitate forms during storage, centrifuge again. Pleural and ascites, cerebrospinal fluid reference implementation. 4. Cell culture supernatant: When detecting secreted components, collect with a sterile tube. Centrifuge for about 20 minutes (2000-3000 rpm). Collect the supernatant carefully. When detecting the components inside the cells, dilute the cell suspension with PBS (PH7.2-7.4), and the cell concentration will reach about 1 million / ml. Through repeated freezing and thawing, the cells are destroyed and the intracellular components are released. Centrifuge for about 20 minutes (2000-3000 rpm). Collect the supernatant carefully. If a precipitate forms during storage, it should be centrifuged again. 5. Organize the specimen: after cutting the specimen, weigh it. Add a certain amount of PBS, PH7.4. Quickly freeze and save with liquid nitrogen for later use. After the specimen melts, it still maintains a temperature of 2-8 ° C. Add a certain amount of PBS (PH7.4) and homogenize the specimen with a manual or homogenizer. Centrifuge for about 20 minutes (2000-3000 rpm). Collect the supernatant carefully. After aliquoting, a portion is to be tested, and the rest is frozen for future use. 6. The specimen should be extracted as soon as possible after collection. The extraction should be carried out according to 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. 7. Samples containing NaN3 cannot be detected because NaN3 inhibits the activity of horseradish peroxidase (HRP). Operating steps Dilution and sample loading of standards: set up 10 wells of the standard on the enzyme-coated plate, add 100 μl of the standard in the first and second wells, and then add the standard in the first and second wells 50μl of diluent, mix well; then take 100μl from the first well and the second well and add them to the third and fourth wells respectively, and then add 50μl of standard diluent to the third and fourth wells respectively, mix well; Then take 50μl each in the third and fourth wells and discard it, then add 50μl each to the fifth and sixth wells, and then add 50ul of the standard dilution solution to the fifth and sixth wells respectively, and mix well; After mixing, take 50μl from the fifth and sixth wells and add them to the seventh and eighth wells respectively. Then add 50μl of the standard dilution solution to the seventh and eighth wells respectively. Take 50μl from the eight wells and add them to the ninth and tenth wells. Then add 50μl of the standard dilution solution to the ninth and tenth wells. After mixing, take 50μl from the ninth and tenth wells and discard. (After dilution, the amount of sample added to each well is 50 μl, and the concentrations are 900 pg / ml, 600 pg / ml, 300 pg / ml, 150 pg / ml, and 75 pg / ml. Sample addition: set blank wells separately (blank control wells without sample and Enzyme label reagent, the rest of the steps are the same), the sample well to be tested. First add 40μl of sample diluent to the sample well of the enzyme plate, and then add 10μl of the sample to be tested (the final dilution of the sample is 5 times). Add the sample and add the sample to the bottom of the well of the microplate, try not to touch the wall of the well, gently shake to mix. Incubation: seal the plate with the sealing plate and incubate at 37 ° C for 30 minutes. Solution: concentrate and wash 20 times Dilute the solution with distilled water 20 times before use. Washing: Carefully remove the sealing film, discard the liquid, spin dry, fill each well with the washing solution, let it stand for 30 seconds and discard, repeat this 5 times, pat dry. Add enzyme : Add 50μl of enzyme-labeled reagent 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 the developer A50μl first to each well, then add the developer B50μl, and gently shake to mix ， Develop at 37 ° C in the dark for 15 minutes. Termination: Add 50μl of stop solution to each well to terminate the reaction Color). Determination: The absorbance (OD value) of each well is measured in sequence with a blank air conditioner at zero, 450 nm wavelength. The measurement should be performed within 15 minutes after the addition of the stop solution. Note: The kit should be removed from the refrigerated environment and equilibrated at room temperature It can be used after 15-30 minutes. If the enzyme-coated plate is not used up after opening, the slat should be stored in a sealed bag. The concentrated washing liquid may crystallize out. When diluted, it can be heated and dissolved in a water bath. Washing does not affect the results. The sampler should be used at each step of sample addition, and its accuracy should be regularly checked to avoid experimental errors. The time of one sample is best controlled within 5 minutes. If there are many specimens, it is recommended to use a volley gun to add Sample. Please make a standard curve at the same time of each measurement, preferably re-well. If the content of the test substance in the specimen is too high (the sample OD value is greater than the OD value of the first well of the standard well), please dilute with the sample diluent first Determine after a certain multiple (n times), and finally multiply by the total dilution multiple (× n × 5) when calculating. The sealing film is only for one-time use to avoid cross-contamination. The substrate should be kept away from light. Strictly follow the instructions Operation, The results of the test must be determined by the reading of the microplate reader. All samples, washing liquid and various wastes should be treated as infectious agents. The components of different batches of this reagent should not be mixed. 10. If there are differences from the English instructions, the English instructions Calculate: Take the concentration of the standard as the abscissa and the OD as the ordinate, draw a standard curve on the graph paper, and find out the corresponding concentration from the standard curve according to the OD value of the sample; then multiply by the dilution factor; or Calculate the linear regression equation of the standard curve using the concentration of the standard and the OD value, substitute the OD value of the sample into the equation, calculate the sample concentration, and multiply it by the dilution factor to obtain the actual concentration of the sample. Kit Performance: 1. Sample The linear regression and expected concentration correlation coefficient R value is more than 0.990. 2. The batch and batch see should be less than 9% and 11% respectively. Detection range: 1.56 pg / ml-100 pg / ml Sensitivity: 0.39 pg / ml Reaction time: 1-5h Required sample volume: 50-100ul Detection wavelength: 450 nm storage conditions and expiration date: 1. Kit storage: 2-8 ℃. 2. Validity: 6 months
Stretch Mark Cream,stretch mark removal cream,lotion for stretch mark,best stretch mark cream
Zchoise Cosmetic Factory , https://www.gzzchoise.com