Rabbit Dehydroepiandrosterone Sulfate (DHEA-S) elisa Kit Instruction Manual Elisa kit Size: 48-well configuration / 96-well configuration Standard diluent: 1.5 ml × 1 bottle Enzyme-labeled Reagent: 3 ml × 1 bottle (48 )/6 ml x 1 bottle (96) [Dehydroepiandrosterone sulphate (DHEA-S) elisa kit] This reagent is for research use only: the concentration of the standard is the abscissa, and the OD is the ordinate. ,Draw a standard curve on the coordinate paper, according to the OD value of the sample from the standard curve to find the corresponding concentration; multiply the dilution multiple; or use the standard concentration and OD value to calculate the standard curve of the linear regression equation, the sample The OD value is substituted into the equation and the sample concentration is calculated and multiplied by the dilution factor, which is the actual concentration of the sample. Kit components: Closure film: 2 sheets (48)/2 sheets (96) Instruction sheet: 1 sealed bag: 1 standard product: 2700 ng/L 0.5 ml x 1 bottle 0.5 ml x 1 bottle 2-8 °C Storage of enzyme Standard plate: 1×48 1×96 2-8°C Sample dilution: 3ml×1 bottle 6 ml×1 bottle 2-8°C Reagent A solution: 3ml×1 bottle 6ml×1 bottle 2 Store the color developer B solution at -8°C: 3ml × 1 bottle 6ml × 1 bottle 2-8°C preservation stop solution: 3ml × 1 bottle 6ml × 1 bottle 2-8°C Concentrate washing solution: (20ml × 20times) × 1 bottle (20 ml × 30 times) × 1 bottle stored at 2-8°C Principle of the experiment: This kit uses a double-antibody sandwich method to determine the level of dehydroepiandrosterone sulphate (DHEA-S) in rabbits. Purified DHEA-S antibody was coated on the microtiter plate to prepare a solid-phase antibody. Dehydroepiandrosterone sulphate (DHEA-S) was added to the micro-wells of the coated monoclonal antibody. ), and then combined with HRP-labeled dehydroepiandrosterone sulphate (DHEA-S) antibody to form an antibody-antigen-enzyme-labeled antibody complex, after thorough washing, plus substrate TMB color. TMB is converted to blue under the catalysis of the HRP enzyme and converted to the final yellow color by the action of acid. The depth of color is positively correlated with dehydroepiandrosterone sulphate (DHEA-S) in the sample. The absorbance (OD value) was measured with a microplate reader at a wavelength of 450 nm, and the concentration of dehydroepiandrosterone sulphate (DHEA-S) in rabbits was calculated from the standard curve. Purpose: This kit is used to determine the content of dehydroepiandrosterone sulphate (DHEA-S) in rabbit serum, plasma and related fluid samples. Service commitment:? Availability: payment and delivery. Free technical advice and guidance during working hours? Please inquire to provide customers with sample testing services to maximize the effectiveness of the experimental results (free agency testing). Preservation conditions and expiration date: Kit storage: 2-8°C | Validity period: 6 months 〠Rabbit dehydroepiandrosterone sulphate (DHEA-S) elisa kit] Sample handling and requirements: 1. Serum: room temperature blood coagulation 10-20 minutes, centrifuge for about 20 minutes (2000-3000 rpm). Carefully collect the supernatant and if sedimentation occurs during storage, centrifuge it again. 2. Plasma: EDTA or sodium citrate should be selected as the anticoagulant according to the requirements of the specimen. After mixing for 10-20 minutes, centrifugation is performed for about 20 minutes (2000-3000 r/min). Carefully collect the supernatant and if any precipitate forms during storage, it should be centrifuged again. 3. Urine: Collect with a sterile tube and centrifuge for about 20 minutes (2000-3000 rpm). Carefully collect the supernatant. If precipitation forms during storage, it should be centrifuged again. Pleural fluid, cerebrospinal fluid reference implementation. 4. Cell culture supernatant: When detecting secretory components, use a sterile tube for collection. Centrifugation for about 20 minutes (2000-3000 rpm). Collect the supernatant carefully. When detecting intracellular components, the cell suspension was diluted with PBS (pH 7.2-7.4) to a cell concentration of about 1 million cells/ml. By repeated freezing and thawing, the cells are destroyed and the intracellular components are released. Centrifugation for about 20 minutes (2000-3000 rpm). Collect the supernatant carefully. If precipitation forms during storage, it should be centrifuged again. 5. Organize the specimen: After cutting the specimen, weigh it. Add a certain amount of PBS, pH 7.4. Liquid nitrogen quickly frozen for use. After the sample melts, it still maintains a temperature of 2-8°C. A certain amount of PBS (pH 7.4) was added and the sample was homogenized by hand or homogenizer. Centrifugation for about 20 minutes (2000-3000 rpm). Collect the supernatant carefully. One part is to be tested after dispensing and the rest is frozen for use. 6. Samples should be extracted as soon as possible after collection. Extraction should be carried out according to the relevant literature. After the extraction, experiments should be carried out as soon as possible. If the test cannot be performed immediately, the specimen can be stored at -20°C but repeated freezing and thawing should be avoided. 7. The sample containing NaN3 cannot be detected because NaN3 inhibits horseradish peroxidase (HRP) activity. Step 1. Dilution and sample addition of standard product: Set 10 standard wells in the enzyme-conjugated plate, add 100 μl of standard in the first and second wells, and add the first and second wells. 50 μl of the standard diluent was mixed; then 100 μl of each of the first and second wells was added to the third well and the fourth well, respectively, and 50 μl of the standard diluent was added to the third and fourth wells respectively. Uniform; then in the third and fourth wells first 50μl discarded each, and then each 50μl were added to the fifth and sixth wells, and then in the fifth and sixth wells were added standard diluent 50ul, mixed After homogenizing, add 50μl from each of the fifth and sixth wells to the seventh and eighth wells after mixing, and then add 50μl of standard diluent to each of the seventh and eighth wells. In the eighth well, 50 μl was added to the ninth and tenth wells, respectively, and 50 μl of the standard diluent was added to the ninth and tenth wells respectively. After mixing, 50 μl of each of the ninth and tenth wells was discarded. (After dilution, the volume of each well was 50 μl, and the concentrations were 1800 ng/L, 1200 ng/L, 600 ng/L, 300 ng/L, and 150 ng/L, respectively). 2. Samples: blank wells (blank control wells without sample and enzyme labeling reagents, and the rest of the steps are the same), sample wells to be tested. 40 μl of sample diluent is added to the well of the sample to be tested on the enzyme-labeled plate, and then 10 μl of the sample to be tested is added (the final dilution of the sample is 5 times). Add the sample to the bottom of the wells of the microtiter plate, try not to touch the wall of the well, mix gently by shaking. 3. Incubation: Seal the plate with a cover plate and incubate at 37°C for 30 minutes. 4. Dosing solution: 30 times (20 times the 48T) diluted washing solution is diluted with distilled water 30 (20 times of 48T) before use. 5. Washing: Carefully remove the sealing film, discard the liquid, spin dry, fill the well with washing liquid, and let it stand for 30 seconds. Discard it. Repeat 5 times and pat dry. 6. Enzyme addition: Add 50 μl of Enzyme Labeling Reagent to each well, except blank wells. 7. Incubation: Operate in the same way as in 3. 8. Washing: Operate with 5. 9. Coloration: Add 50 μl of color reagent A to each well, add 50 μl of color reagent B, mix gently, and avoid color at 37°C 15 minutes. 10. Termination: Add 50 μl of stop solution to each well and stop the reaction (in this case the blue turns to yellow). 11. Measurement: The absorbance (OD value) of each well was measured sequentially with a blank air conditioner at zero wavelength of 450 nm. The measurement should be performed within 15 minutes after adding the stop solution. Dehydroepiandrosterone sulphate (DHEA-S) elisa kit Note: 1. The kit should be removed from a refrigerated environment and allowed to equilibrate for 15-30 minutes at room temperature. If the ELISA plate is unopened after opening, the slats should be stored in a sealed bag. 2. The concentrated washing solution may have crystals precipitated, and may be heated and dissolved in a water bath during dilution. The washing does not affect the results. 3. The sampler should be used for each step, and the accuracy of the sampler should always be calibrated to avoid experimental errors. It is best to control the injection time within 5 minutes at a time. If the number of specimens is large, it is recommended to use a sample of guns. 4. Please make a standard curve at the same time for each measurement. It is best to make a duplicate hole. If the content of the test substance in the sample is too high (the OD value of the sample is larger than the OD value of the first hole of the standard product hole), please dilute a certain multiple (n times) with the sample diluent and then measure. Calculate the total dilution by multiplying Multiple (×n×5). 5. Sealing film is limited to one-time use to avoid cross-contamination. 6. Protect the substrate from light. 7. In strict accordance with the instructions of the operation, the test results must be determined by the microplate reader. 8. All samples, washing liquids and all kinds of waste should be handled as infectious agents. 9. The lot number components of this reagent must not be mixed. 10. If there is a difference with the English manual, the English manual shall prevail. Performance of the kit: 1. The correlation coefficient between the linear regression of the sample and the expected concentration is 0.95 or more. 2. Within the batch and batch should be less than 9% and 11% respectively Detection range: 0.2IU/L - 6IU/L