The effect of coating on the ELISA kit experiment! -Huaqiang Electronic Network

The Impact of Coating on ELISA Kit Experiments! When performing experiments with an ELISA kit, there are numerous factors to consider. Biologists often highlight issues such as incubation time, color development, and coating. Among these, the coating process is one of the most critical steps, directly affecting the accuracy and reliability of the results. How to Perform Coating? Coating involves fixing an antigen or antibody onto a solid-phase carrier. This is typically done through physical adsorption, where proteins bind to polystyrene surfaces based on hydrophobic interactions. This non-specific binding is influenced by the protein’s molecular weight, isoelectric point, and concentration. Larger macromolecular proteins usually have more hydrophobic regions, making them easier to adsorb onto the solid support. However, smaller proteins may not bind effectively. In such cases, indirect methods like affinity chromatography can be used to improve the purity of the coated antigen. For antigens with high impurity levels, a capture coating method is also effective. Another common approach is using the avidin-biotin system. Avidin is first coated onto the surface, followed by biotinylated DNA. This method ensures uniform and stable binding, and it has been widely applied in quantitative detection of various antigens. For lipid substances that cannot bind directly to the solid phase, they can be dissolved in organic solvents like ethanol, added to the ELISA plate, and then dried after evaporation. Advantages of Proper Coating: Proper coating improves the specificity and sensitivity of the ELISA test, enhances repeatability, and reduces the amount of antigen needed—often by as much as 1/10 or even 1/100 compared to direct coating. Coating Antigens: There are three main types of antigens used in ELISA: natural antigens, recombinant antigens, and synthetic peptide antigens. Synthetic peptides are artificially made fragments of antigenic determinants. They are highly pure and specific but have low molecular weight, which makes direct adsorption difficult. In such cases, conjugates are used to indirectly bind the antigen to the solid support. Coating Antibodies: IgG antibodies have strong adsorption capabilities to polystyrene, particularly through their Fc region. This leaves the antigen-binding site exposed, which is ideal for ELISA applications. When using antiserum or cell culture supernatants containing monoclonal antibodies, it's important to remove any hybrid antibodies to ensure test specificity. Monoclonal antibodies from ascites fluid are often more concentrated and specific, so they don’t require additional purification before use. In some cases, combining multiple monoclonal antibodies can lead to better performance. Optimal Coating Conditions: Common coating buffers include: - pH 9.6 carbonate buffer - pH 7.2 phosphate buffer - pH 7–8 Tris-HCl buffer After adding the coating solution, it is usually incubated at 4–8°C overnight or at 37°C for 2 hours. The optimal concentration of the coating antigen varies depending on the carrier material and the nature of the protein. Typical concentrations range from 10 ng/mL to 20 µg/mL. Each batch should be tested to determine the best conditions for optimal performance.

Rocker Switch

Features

â—† Wide Application

Widely used for various kinds of electrical products, instrument, car, boat, household appliances such as lights, water dispenser, treadmill, coffee pot, speaker, electric car, motorcycle, TV, massage machine etc.

â—† Easy to install and use

3 PIN on-off rocker switch with SPST design, simple installation, freely turn on or off the load which you want to control.

â—† High Operating Life

Made of high quality polyamide eP(Nylon PA66) material, this sturdy mini boat rocker switch is born for anti-corrosion,anti-acid and high resistant with silver terminals.100,000 times of ON/OFF operating life span.

Rocker Switch,Waterproof Rocker Switch,Custom Rocker Switches,Mini Rocker Switch

Ningbo Jialin Electronics Co.,Ltd , https://www.donghai-switch.com