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Zebrafish Advanced glycation End Products (AGEs)ELISA Manual
AGES is the English abbreviation for glycosylation end products, which are the end products of non-enzymatic glycosylation reactions (Maillard reactions). It refers to the stable covalent admixture generated by the spontaneous reaction of macromolecules such as proteins, lipids or nucleic acids with glucose or other reducing monosaccharides without the participation of enzymes. This kit was used for the determination of advanced glycation end products (AGEs) in serum, plasma and related fluid samples of zebrafish. So what are the precautions in the ELISA experiment? Hengyuan Biological Xiaobian summarizes the precautions for the operation of the ELISA experiment.
First, experimental principle
This kit was used to determine the levels of advanced glycation end products (AGEs) in zebrafish samples by double antibody sandwich method. Purified zebrafish advanced glycation end products (AGEs) antibodies were coated with microporous plates to prepare solid phase antibodies, and advanced glycation end products (AGEs) were added to the micropores coated with mab successively, and then combined with advanced glycation end products (AGEs) antibodies labeled by HRP to form antibody-antibody-enzymic antibody complex. After thorough washing, add substrate TMB for color development. The TMB is catalyzed by HRP enzymes to turn blue and converted to the final yellow color under the action of acid. The color depth was positively correlated with advanced glycation end products (AGEs) in the sample. The absorbance (OD) was measured at 450nm wavelength by enzymoleter. The concentrations of advanced glycation end products (AGEs) in zebrafish samples were calculated by standard curve.
2. Specimen requirements
1. The specimen should be extracted as soon as possible after collection, and the extraction should be carried out according to relevant literature, and the experiment should be conducted as soon as possible after extraction. If the test cannot be carried out immediately, the specimen can be stored at -20℃, but repeated freezing and thawing should be avoided
2. Samples containing NaN3 cannot be detected because NaN3 inhibits horseradish peroxidase (HRP) activity.
Third, precautions
1. After the kit is removed from the refrigerated environment, it should be balanced at room temperature for 15-30 minutes before use. If the enzyme-labeled kit is not used up after being opened by the plate, the strip should be stored in a sealed bag.
2. The concentrated washing solution may crystallize out. When diluted, it can be heated in the water bath to help dissolve, and the result will not be affected during washing.
3. The sample feeder should be used at each step, and its accuracy should be checked frequently to avoid test errors. It is best to control the sampling time within 5 minutes. If the number of specimens is large, it is recommended to use volley sampling.
4. Please do the standard curve at the same time each measurement, preferably do the double hole. If the content of the substance to be measured in the sample is too high (the OD value of the sample is greater than the OD value of the first hole of the standard product), please dilute the sample diluent with a certain number (n times) before measuring, and then multiply the total dilution times (×n×5) when calculating.
5. The sealing film is for one-time use only to avoid cross contamination.
6. Keep the substrate away from light.
Seven. The operation is carried out in strict accordance with the instructions, and the test result must be determined by the reading of the enzyme label instrument.
8. All samples, washing fluids and various wastes should be treated as infectious agents.
9. Different batch number components of this reagent should not be mixed.
The complete version of the manual can consult the salesman to obtain oh, Hengyuan biological has a group of skilled, experienced technical engineers, after years of accumulation, the kit now covers a lot of species, more than thousands of indicators, with fast, simple, accurate characteristics, to meet the needs of the majority of universities and researchers, improve the efficiency of researchers, mature and stable quality, It has won the recognition of many scientific research institutions, perfect inventory and supply system and efficient and stable technical guidance to ensure that products can be supplied in stock. Look forward to your consultation.
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