MT ELISA Kit
MT ELISA Kit
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Product Details
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Product Specification
| protein | MT | |||||||||||||||||||||||||||||||||
| Usage |
Self-prepared test equipment required for the experiment: 1 , plate reader ( 450nm ） 2 , high-precision sampler and gun head: 0.5-10uL 、 5-50uL 、 20-200uL 、 200-1000uL 3 、 37℃ Incubator 4 Distilled water or deionized water, Sample handling and requirements: Serum: Whole blood samples collected in serum separation tubes were placed at room temperature 2 Hour or 4℃ Overnight, then 1000×g Centrifugation 20 Minutes, take the supernatant, or place the supernatant in -20℃ Or -80℃ Store, but repeated freezing and thawing should be avoided. Plasma: use EDTA Or heparin as an anticoagulant to collect specimens, and collect the specimens after collection 30 Within minutes 2-8℃1000×g Centrifugation 15 Minutes, take the supernatant to detect, or place the supernatant in -20℃ Or -80℃℃ Store, but repeated freezing and thawing should be avoided. Other biological fluids: 1000xg Centrifugation 20 Minutes, take the supernatant to detect. Preparations before testing: 1 , please advance 10 Minutes remove the kit from the refrigerator and equilibrate to room temperature. 2 , Standard gradient working solution preparation : join 1mL Universal Diluent into Lyophilized Standard , let stand 15 Minutes until it is completely dissolved and then gently mix (the concentration is 500pg/mL ） And then according to the following concentrations: 500pg/mL 、 250pg/mL 、 125pg/mL 、 62.5pg/mL 、 31.25pg/mL 、 15.625pg/mL 、 7.8125pg/mL 、 0pg/mL The dilution was performed. Double dilution method: Take 7 branch EP Tubes, each tube is added 500uL Universal diluent ,500pg/mL Pipette from the standard working solution 500uL To the first EP Mix evenly in a tube 250pg/mL Standard Working Solution , according to this step, absorb and mix evenly in turn. The last tube is directly used as a blank hole, There is no need to draw liquid from the penultimate tube, as shown in the figure below.  3 、 Biotin- Preparation of antibody working solution : Before use 15 Minutes will concentrate Biotin- Antibody to 1000×g Centrifugation 1 Minutes, with a universal diluent 100× concentrate Biotin- The antibody is diluted into 1× Working concentration (ex: 10uL Concentrate +990uL Universal Diluent) , used on the same day. 4 、 Preparation of enzyme conjugate working solution : Before use 15 Minutes will 100x Concentrated enzyme conjugate in 1000×g Centrifugation 1 Minutes, with a universal diluent 100× concentrate HRP The enzyme conjugate is diluted into 1× Working concentration (ex: 10uL Concentrate +990uL Universal Diluent), for same day use. 5 、 1× Wash liquid preparation: Take 10mL 20× Wash liquid to 190mL In distilled water (the concentrated washing liquid taken out of the refrigerator may have crystals, which is a normal phenomenon. It can be left at room temperature and prepared after the crystals are completely dissolved). Operation steps: 1 Equilibration from room temperature 10 After minutes, remove the required slats from the aluminum foil bag, and seal the remaining slats with ziplock bag and put them back 4℃ 。 2 , Adding samples: respectively add samples or different concentration standards according to 50ul Each well is added to the corresponding well, and the blank well is added 50uL Universal dilution followed by each well 50uL Biotin- Antibody working fluid. After covering the sealing film 37℃ Incubation 1 Hours. (Recommendation: minimum dilution of sample to be tested with universal diluent 1 Then add enzyme labeled in-plate test, so as to reduce the error of matrix effect on test results , influence, and finally calculate the sample concentration by multiplying it by the corresponding dilution factor. It is recommended to set up double wells for all samples and standards to be tested during testing). 3 Plate washing: discard the liquid and add to each well 300uL 1x Wash liquid, stand 1 Minutes, throw off the washing liquid, pat dry on absorbent paper, and repeat washing the plate 3 Times (the plate can also be washed with a plate washing machine). 4 Adding enzyme conjugate working solution Enzyme conjugate working solution is added to each well 100uL , after covering the sealing film 37℃ Incubation 30 Minutes. 5 、 Washing plate : Discard liquid according to step 3 Washing Method , wash plate 5 Times. 6 Substrate addition: substrate is added per well ( TMB ） 90uL Covered with a sealing film, 37℃ Incubation protected from light 15 Minutes. 7 Add stop solution: take out the enzyme label plate and directly add stop solution to each well 50uL , immediately in 450nm Wavelength measurement of each well OD Value. Calculation of experimental results: Result judgment: 1 , calculate the average of the standard and sample replica well OD Value and subtract the blank hole's OD Values as correction values. Taking concentration as the abscissa, OD Value is ordinate , draw the standard curve of the four-parameter logic function on the double logarithmic coordinate paper. 2 If the sample OD If the value is higher than the upper limit of the standard curve, the test should be retested after appropriate dilution and multiplied by the corresponding dilution factor when calculating the sample concentration.
 Kit Performance: 1 Repeatability: the coefficient of variation in the plate is less than 10% , the interplate coefficient of variation is less than 10% 。 2 Recovery rate: adding to selected healthy serum and plasma 3 Of different concentration levels MT , calculate the recovery.
3 , linear dilution: respectively in the selected 4 Healthy serum and plasma are added with high concentration MT , linearity was assessed by dilution within the standard curve kinetic range.
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| Theory | This kit uses competitive enzyme-linked immunosorbent assay (ELISA). The sample, standard, Biotin-labeled antibody, and HRP enzyme conjugate were sequentially added to the microwells pre-coated with the universal species Melatonin (MT) antigen, incubated and washed in the middle, and the substrate TMB was used to develop color. The TMB was converted to blue under the catalysis of peroxidase (HRP), and to the final yellow under the action of acid. The depth of color was positively correlated with the universal species Melatonin (MT) in the samples. The absorbance (OD value) was measured with a microplate reader at a wavelength of 450nm, and the sample concentration was calculated. | |||||||||||||||||||||||||||||||||
| Source | General purpose | |||||||||||||||||||||||||||||||||
| Synonym | Melatonin ELISA Kit | |||||||||||||||||||||||||||||||||
| Detection Type | Competition Law | |||||||||||||||||||||||||||||||||
| Composition |
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| Background | melatonin, or melatonin or melatonin, is chemically known as N-acetyl-5-methoxytryptamine. In universal species, serotonin (also known as serotonin) is converted to N-acetyl-5-hydroxytryptamine by reaction with acetyl-CoA catalyzed by N-acetyltransferase, which is then methylated to melatonin by S-adenosylmethionine catalyzed by acetyl-serotonin O-methyltransferase. The side effects of melatonin supplements are minimal when given in small doses, short courses. They may include somnolence (drowsiness), headache, nausea, diarrhea, abnormal dreams, irritability, nervousness, restlessness, insomnia, anxiety, migraine, drowsiness, psychomotor hyperactivity, dizziness, high blood pressure, abdominal pain, heartburn, mouth ulcers, dry mouth, hyperbilirubinemia. Dermatitis, night sweats, itching, rash, dry skin, pain in extremities, menopausal symptoms, chest pain, glycosuria (sugar in urine), proteinuria (protein in urine), abnormal liver function tests, weight gain, fatigue, mood swings, aggression and a feeling of hangover. It is not recommended for use during pregnancy or breastfeeding or in generic species with liver disease. | |||||||||||||||||||||||||||||||||
| General Notes | 1. Carry out incubation in strict accordance with the specified time and temperature to ensure accurate results. All reagents must reach room temperature 20-25 °C prior to use. Store reagents in refrigeration immediately after use. 2. Incorrect plate washing may lead to inaccurate results. Make sure to drain the liquid from the wells as much as possible before adding the substrate. Do not allow the wells to dry out during incubation. 3. Eliminate the residual liquid and fingerprints at the bottom of the plate, otherwise it will affect the OD value. 4. The substrate color development solution should be colorless or very light in color, and the substrate solution that has turned blue cannot be used. 5. Avoid cross-contamination of reagents and specimens to avoid wrong results. 6. Avoid direct exposure to strong light during storage and incubation. 7. Any reaction reagent cannot come into contact with the bleaching solvent or the strong gas emitted by the bleaching solvent. Any bleaching component will destroy the biological activity of the reaction reagents in the kit. 8. Expired products cannot be used, and components with different item numbers and batch numbers cannot be mixed. 9. Recombinant proteins from sources other than the kit may not match the antibodies in this kit and are not recognized. 10. If the disease may be spread, all samples should be managed well, and the samples and testing devices should be handled according to the prescribed procedures. |
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| Storage Temp. | Unopened kit, stored at 4 °C, shelf life 6 months. | |||||||||||||||||||||||||||||||||
| Test Range | 7.8-500pg/mL | |||||||||||||||||||||||||||||||||
| Applications | Serum, plasma and other biological fluids |
