Chicken insulin (INS) enzyme-linked immunoassay kit instructions

Chicken Insulin (INS) ELISA Kit Instruction Manual This kit is for research use only.
Detection range: 96T
This kit is used to determine insulin (I) levels in chicken serum, plasma and related fluid samples.
Principle of the experiment This kit uses a double antibody sandwich method to determine the level of chicken insulin (I) in the sample. A solid-phase antibody is prepared by coating a microtiter plate with a purified chicken insulin (I) antibody, insulin (I) is sequentially added to the micropores of the coated monoclonal antibody, and the antibody is bound to the HRP-labeled insulin (I) antibody to form an antibody. - Antigen-enzyme-labelled antibody complexes, after thorough washing, were developed with substrate TMB. 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 insulin (I) in the sample. The absorbance (OD value) was measured with a microplate reader at a wavelength of 450 nm, and the chicken insulin (I) concentration in the sample was calculated using a standard curve.
Specimen requirements:
1. Serum: Use tubes containing no pyrogens and endotoxins to avoid any stimulation of cells during the operation. After collecting blood, the serum and red blood cells are quickly and carefully separated by centrifugation at 3000 rpm for 10 minutes.
2. Plasma: Anticoagulation with EDTA, citrate, or heparin. Centrifuge at 3000 rpm for 30 minutes to take the supernatant.
3. Cell supernatant: Centrifuge at 3000 rpm for 10 minutes to remove particles and polymer.
4. Homogenize the tissue: Add the tissue to the appropriate amount of saline and mash it. Centrifuge at 3000 rpm for 10 minutes to take the supernatant.
5. Save: If the sample is not collected in time after collection, please dispense it once in one dose and store it at -20°C to avoid repeated freezing and thawing. Thaw at room temperature and ensure that the sample is fully thawed evenly.
Steps
1. Remove the required slats from the aluminum foil bag after equilibration at room temperature for 20 minutes. Seal the remaining slats back to 4°C in a ziplock bag.
2. Set standard product wells, sample wells, and blank wells. Blank wells do not add anything; standard wells add 50 μL of different concentrations of the standard;
3. Add 10 μL of test sample to the sample hole to be tested, and add 40 μL of sample diluent;
4. Followed by adding horseradish peroxidase (HRP)-labeled detection antibody (50μL) to standard wells and sample wells (without blank wells). Seal the reaction wells with a sealing membrane. 37°C water bath or constant temperature 60min.
5. Discard the liquid and pat dry on absorbent paper. Fill each well with washing liquid and let it stand for 1 min. Scour the washing solution and pat dry on the blotting paper. Repeat washing for 5 times (washing with plate washer is also available).
6. Add 50 μL of substrate A and B to all wells and incubate at 37°C for 15 min in the dark.
7. Add 50 μL of stop solution to all wells. Determine the OD of each well at a wavelength of 450 nm within 15 minutes.
The company's elisa kits can provide free agency service. Customers can provide their own proxy samples and provide customers with the required raw data within seven working days.
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