Hengyuan to share the choice of four ELISA experimental conditions - Database & Sql Blog Articles

EL-C1600N100013-B

Heng Jiaxing AO3401

The outer diameter of the probe domestically produced P125-Q2 four-pronged head nickel-plated needle is 2.02 needle

ELISA experiments are highly dependent on the selection of proper experimental conditions. Today, Shanghai Hengyuan will walk you through the key factors that influence the success of an ELISA test. We'll cover the choice of solid-phase carriers, the coating antibody or antigen, the working concentration of the enzyme-labeled antibody, and the selection of the substrate and hydrogen donor. Let's take a closer look at each one:

1. Selection of Solid-Phase Carriers
Various materials can be used as solid-phase carriers, such as polystyrene, polyvinyl chloride, and cellulose. These can come in different forms like plates, tubes, or beads. The most commonly used today is a 40-well polystyrene plate. Before using any carrier, it’s essential to perform a preliminary screening. Coat the surface with the same amount of antigen under identical conditions and check for uniform color development and good adsorption performance.

2. Choice of Coating Antibody (or Antigen)
The antibody or antigen must be of high purity, and the pH should generally be between 9.0 and 9.6. Factors like temperature, time, and protein concentration also play a role. Typically, coating is done at 4°C for 18–24 hours. To determine the optimal concentration, different protein concentrations (e.g., 0.1, 1.0, 10 μg/ml) are tested. The concentration that gives the highest OD value while using the least amount of protein is usually chosen, typically between 1–10 μg/ml for most proteins.

3. Working Concentration of Enzyme-Labeled Antibody
Start by titrating the initial titer using direct ELISA. Then, fix other conditions or use the “square matrix method” (where the coating, sample, and enzyme-labeled antibody are all tested at different dilutions) to accurately determine the optimal working concentration in the actual experimental system.

4. Selection of Substrate and Hydrogen Donors
The hydrogen donor should be cost-effective, safe, and produce a clear color change without being colored itself. Some options, like OPD, may have potential carcinogenic effects and require careful handling. Safer alternatives include TMB and ABTS, which are widely used today. After the reaction has proceeded for about 10–30 minutes, the reaction is usually stopped by adding a strong acid or base. It’s important to prepare the substrate fresh before use, especially hydrogen peroxide (H₂O₂).

Shanghai Hengyuan continues to offer cutting-edge ELISA technologies. Our company provides high-quality ELISA kits, both imported and domestic, suitable for human, rat, and mouse samples. We guarantee quality and competitive pricing. If you're interested in learning more or purchasing our products, feel free to contact our sales team for assistance.