Experimental principle and operation procedure of monkey activator of transcription-5 (ATF-5) enzyme-linked immunoassay kit.
1. Experimental principle.
The Monkey Activator of Transcription-5 (ATF-5) ELISA Kit uses indirect enzyme-linked immunoassay (ELISA) to capture and detect ATF-5 in the sample to be tested by using the specific binding reaction of antigen and antibody. The ATF-5 antibody is coated on a microplate and the ATF-5 in the sample binds to the coated antibody, and the enzyme-labeled ATF-5 antibody is added to form an immune complex. After washing, the substrate chromogenic solution is added and the color is developed by an enzyme-catalyzed reaction. Finally, the optical density value was determined by the microplate reader, and the concentration of ATF-5 in the sample to be measured was calculated.
2. Steps.
1.Prepare reagents and materials: monkey activator of transcription-5 (ATF-5) enzyme-linked immunoassay kit, microplate plate, washing solution, sample diluent, pipette, lab coat, gloves, mask, recording pen, etc.
2.Sample Processing: According to the requirements of the kit instructions, the sample to be tested is appropriately diluted and an equal amount of sample dilution is added.
3.Dispensing: Use a pipette to add the diluted sample to the appropriate wells on the plate and follow the dosing order and amount required by the kit instructions.
4.Incubation: The microplate plate is sealed and incubated at 37 for a certain period of time to allow the antigen and antibody to fully bind.
5.Washing: Wash the plate for a certain number of times according to the requirements of the kit instructions to remove unbound antigens, antibodies and impurities.
6.Enzyme-labeled antibody: Use a pipette to add the enzyme-labeled antibody to the appropriate well on the plate.
7.Re-incubation: The plate is sealed and incubated again at 37 for a certain period of time to allow the enzyme-labeled antibody to bind to the antigen-antibody complex.
8.Washing: Wash the plate again as required by the kit instructions for a certain number of times to remove unbound enzyme-labeled antibodies and impurities.
9.Add substrate chromogen: Use a pipette to add the substrate chromogen to the appropriate wells on the plate plate.
10.Color development: The plate is left at room temperature for a certain amount of time and a noticeable color change is observed.
11.Stop the reaction: Use a pipette to add the stop solution to the appropriate well on the plate to stop the chromogenic reaction.
12.Determination of optical density value: The optical density value of each well is determined using a microplate reader, and the data is recorded.
13.Data processing and analysis of results: Convert the optical density value to the concentration of ATF-5 according to the formula or graph provided in the kit instructions. Data analysis was carried out to draw experimental conclusions.
It should be noted that the operation procedure of this experiment is for reference only, and the specific operation should be carried out according to the requirements of the kit manual. At the same time, it is necessary to keep the laboratory clean and hygienic during the experiment, comply with the laboratory safety regulations, and prevent cross-contamination and accidents.