Fish tight junction protein detection kit range and experimental principle method

Fish tight junction protein detection range and experimental principle method

Fish tight junction protein (Occludin) ELISA test kit
English name: Fish Occludin ELISA Kit
Type of experiment: Sandwich method detection range: 0.625 ng/mL – 20 ng/mL
Minimum detection limit: the minimum detection concentration is less than 0.1 ng/mL
Applications: Serum, plasma, tissue homogenate, cell culture supernatant or other related fluids (this product is for laboratory research and non-clinical use only)
Explanation <br> Due to the existing conditions and scientific and technical level, it is not possible to comprehensively identify and analyze all the raw materials provided by all suppliers. This product may have certain quality and technical risks.
  • The final experimental results are closely related to the effectiveness of the reagents, the experimenter's related operations, and the experimental environment at the time. Be sure to prepare sufficient specimen backups.
  • Different batches of the same product may have a slight difference, such as: detection limit, sensitivity and color development time, etc., please carry out the experimental operation according to the instructions in the kit. The electronic version of the website is for reference only.
  • Only the reagents used in this kit can be used to ensure the test results. Do not mix products from other manufacturers. The best test results will only be obtained if the experimental instructions of this kit are strictly followed.
  • The company is only responsible for the kit itself, and is not responsible for the sample consumption caused by the use of the kit. Please fully consider the possible usage of the sample before use, and reserve sufficient samples.
  • Tissue homogenates or cell extracts prepared using chemical lysates may cause bias in ELISA results due to the introduction of certain chemicals.
  • If the sample is a cell culture supernatant, there are many factors that interfere with such samples, such as cell status, cell number, sampling time, etc., so there may be cases where detection is not possible.
  • Certain natural or recombinant proteins, including prokaryotic and eukaryotic recombinant proteins, may not be detected because they do not match the detection and capture antibodies used in this product.
Experimental principle The kit uses a double antibody one-step sandwich enzyme-linked immunosorbent assay (ELISA). To the coated microwells pre-coated with the fish tight junction protein (Occludin) capture antibody, the test sample, the standard, the HRP-labeled detection antibody were sequentially added, and the cells were washed and thoroughly washed. Using the substrate TMB to develop color, TMB is converted to blue under the catalysis of peroxidase and converted to the final yellow color by the action of an acid. The color depth is positively correlated with the fish tight junction protein (Occludin) in the sample. The absorbance (OD value) was measured at 450 nm using a microplate reader to calculate the sample concentration.

Steps
  1. The required slats were taken out from the aluminum foil pouch after equilibrating for 20 min at room temperature, and the remaining slats were sealed back to 4 ° C with a ziplock bag.
  2. Set standard and sample wells, standard wells with different concentrations of standard 50μL;
  3. Add 50 μL of the sample to be tested to the sample well; blank holes are not added.
  4. In addition to the blank wells, 100 μL of horseradish peroxidase (HRP)-labeled detection antibody was added to each well of the standard well and the sample well, and the reaction well was sealed with a sealing plate, and incubated at 37 ° C in a water bath or incubator for 60 min.
  5. Discard the liquid, pat dry on the absorbent paper, fill each well with the washing solution (350 μL), let stand for 1 min, remove the washing solution, pat dry on the absorbent paper, and repeat the washing 5 times (can also be washed with a washing machine) .
  6. 50 μL of each of the substrates A and B was added to each well, and incubated at 37 ° C for 15 min in the dark.
  7. 50 μL of the stop solution was added to each well, and the OD value of each well was measured at a wavelength of 450 nm within 15 min.

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