Overview[1]
Human lipoteichoic acid (LTA) ELISA kit is used to determine the lipoteichoic acid (LTA) content in human serum, plasma and related liquid samples. Experimental principle: The double-antibody sandwich method is used to determine the level of human lipoteichoic acid (LTA) in specimens. Use purified human lipoteichoic acid (LTA) antibody to coat the microwell plate to make a solid-phase antibody. Add lipoteichoic acid (LTA) sequentially to the microwells coated with the monoclonal antibody, and then mix it with HRP-labeled lipoteichoic acid. The acid (LTA) antibody combines with the anti-fire hydraulic oil to form an antibody-antigen-enzyme-labeled antibody complex. After thorough washing, the substrate TMB is added for color development. TMB is converted into blue under the catalysis of HRP enzyme and into the final yellow under the action of acid. The depth of color is positively correlated with lipoteichoic acid (LTA) in the sample. Use a microplate reader to measure the absorbance (OD value) at a wavelength of 450 nm, and calculate the concentration of human lipoteichoic acid (LTA) in the sample through the standard curve.
Operation steps[1]
1. Dilution of standards
2. Add samples: Set up blank holes (no samples and enzyme-labeled reagents are added to the blank control holes, and the remaining steps are the same), standard holes, and sample holes to be tested. Accurately add 50 μl of the standard on the enzyme-labeled plate, add 40 μl of sample diluent to the well of the sample to be tested, and then add 10 μl of the sample to be tested (the final dilution of the sample is 5 times). Add the sample to the bottom of the well of the enzyme plate, try not to touch the wall of the well, and shake gently to mix.
3. Incubation: Seal the plate with sealing film and incubate at 37°C for 30 minutes.
4. Solution preparation: Dilute the 30-fold concentrated washing solution with distilled water 30-fold and set aside.
5. Washing: Carefully remove the sealing film, discard the liquid, spin dry, fill each well with washing liquid, let it stand for 30 seconds and then discard, repeat this 5 times, and pat dry.
6. Add enzyme: Add 50 μl of enzyme label reagent to each well, except blank wells.
7. Incubation: The operation is the same as 3.
8. Washing: The operation is the same as step 5.
9. Color development: First add 50 μl of chromogen A to each well, then add 50 μl of chromogen B, shake gently to mix, and develop color for 15 minutes at 37°C in the dark.
10. Termination: Add 50 μl of stop solution to each well to terminate the reaction (blue turns to yellow immediately).
11. Measurement: Measure the absorbance (OD value) of each well with a blank air conditioner zero and a wavelength of 450nm. The measurement should be performed within 15 minutes after adding the stop solution.
