EL-C1600N100013-B
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Kaixin micro test
Test probe P100-M3
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When conducting experiments with ELISA kits, there are several critical factors to consider. Biologists often highlight issues like incubation time, color development, and most importantly, the coating process. The coating step is particularly crucial as it directly affects the accuracy and reliability of the experiment.
How Coating Works
Coating involves fixing an antigen or antibody onto a solid-phase support. This is typically achieved through physical adsorption between the protein and the polystyrene surface. The binding occurs due to interactions between hydrophobic groups in the protein and those on the solid support. This non-specific adsorption can be influenced by factors such as the protein's molecular weight, isoelectric point, and concentration.
Larger macromolecular proteins tend to have more hydrophobic regions, making them easier to bind to the solid phase. Smaller proteins may not adsorb as effectively, but this can be improved by using indirect methods, such as affinity chromatography. In cases where the antigen contains impurities, a capture coating technique can enhance purity and improve experimental outcomes.
Another common method involves using streptavidin-biotin systems. Streptavidin is first coated onto the plate, followed by biotinylated molecules. This approach ensures a uniform and stable coating, and has been widely used for quantifying various antigens. For lipid-based substances that do not bind well to the solid phase, they can be dissolved in organic solvents like ethanol, added to the ELISA plate, and then dried after evaporation.
Benefits of Proper Coating
A well-executed coating improves the specificity and sensitivity of the ELISA test, while also ensuring good reproducibility. It also reduces the amount of antigen needed—often just 1/10 or even 1/100 of what is required for direct coating.
Coating Antigens
There are three main types of antigens used: natural, recombinant, and synthetic peptides. Synthetic peptide antigens are chemically synthesized fragments of larger antigenic determinants. These usually contain only one epitope, offering high purity and specificity. However, their small size makes them less likely to bind directly to the solid phase. In such cases, conjugation techniques are used to attach them to the surface.
Coating Antibodies
Immunoglobulin G (IgG) has a strong affinity for polystyrene surfaces, binding primarily through its Fc region. This leaves the antigen-binding site exposed, which is ideal for ELISA applications. When using antiserum or monoclonal antibody cultures, it's important to remove any interfering antibodies before use to maintain test specificity.
Monoclonal antibodies from ascites fluid are highly concentrated and specific, so they can often be used without further purification. In some cases, mixing different monoclonal antibodies can yield better results.
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, the plate is usually incubated at 4–8°C overnight or at 37°C for 2 hours. The optimal coating concentration varies depending on the carrier material and antigen type. Typical ranges are between 10 ng/ml and 20 µg/ml. Each batch should be tested to determine the best conditions for the specific experiment.
Description
-Contact Resistance:≤50mΩ
-Insulation Resistance:≥100mΩ
-Dielectric Strength:1,500V,
-1min Electronic Life:10,000 cycles
-Operating temperature:T120
-Rating current/voltage:6A 250V AC
Features
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â—† Micro contact gap,High speed operation,High sensitirity,Micro operatizon travel.
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