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What is the relationship between the latest ELISA tips and the test reference?
Yesterday afternoon, in response to a customer's question, Mr. Wang Hengyuan, our technical commissioner, provided a detailed phone explanation regarding the relationship between ELISA and reference standards. He emphasized that quantitative ELISA kits must include reference standards for constructing standard curves, typically consisting of 4–5 concentrations that span the detectable range. These standards are usually prepared in buffers containing protein protectants and preservatives to ensure stability and accuracy.
During the discussion, our technical team highlighted key points about the importance of reference materials:
First, the positive control. This is a critical component used to validate the experiment and serve as a benchmark for interpreting results. The composition of the reference material, especially the negative control, should closely resemble that of the test sample. For instance, when testing human serum, it's best to use human serum as the control. Normal human serum can generate varying levels of background depending on the ELISA format. At Shanghai Hengyuan Biotech Co., Ltd., we offer high-quality ELISA kits. However, obtaining large quantities of normal human serum is challenging, which is why many foreign ELISA kits use recalcified human plasma—plasma with calcium ions added to induce clotting, and then the clot is removed to obtain a liquid similar to serum.
Second, the negative control. It should ideally not contain the target substance being tested. For example, a negative control for HBsAg detection should not have HBsAg and preferably also be negative for anti-HBs. The positive control is usually prepared in a buffer containing a protein stabilizer, with a known amount of the target analyte added. The concentration of this analyte should be clearly indicated in the kit’s instructions and should match the sensitivity of the reagent. By comparing the absorbance of the control with that of the sample, you can estimate the concentration of the target substance in the specimen. For instance, the sensitivity of an HBsAg ELISA kit is around 0.5 ng/ml, and the positive control typically contains about 10 ng/ml of HBsAg. Preservatives and antibiotics are often included in the controls to extend shelf life.
**How to Choose an ELISA Kit**
1. Determine which proteins you want to detect.
2. Identify which species have the highest homology with the gene encoding the protein.
3. Look for kits available for those species.
4. Check with the supplier to learn which type of antibody is used in the kit.
5. Make your own decision based on the information gathered.
Choosing the right ELISA kit is crucial for accurate and reliable results. Always consider the specific needs of your experiment and consult with experts if needed. Whether you're working with human samples or other biological materials, understanding the role of reference standards and controls is essential for successful ELISA performance.