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How to determine whether the results of ELISA experiments are accurate - Database & Sql Blog Articles
ELISA experiments have been completed, and the accuracy of the data obtained depends on a variety of factors. Whether the results meet your expectations is influenced by several key conditions. Today, Shanghai Jinma will share with you the essential elements that contribute to successful ELISA testing.
First and foremost, high-quality reagents are crucial. High-quality reagents, combined with proper instrumentation and correct operation, are fundamental to achieving accurate and reliable ELISA results. The procedure for ELISA can vary depending on the type of solid-phase carrier used. In domestic medical testing, microtiter plates are commonly used. This article will focus on plate-based ELISA, as well as bead, tube, and magnetic bead ELISA methods. When using foreign reagents, it's important to use specialized instruments and strictly follow the operating procedures to ensure accurate outcomes.
Secondly, sample selection plays a significant role in ELISA testing. A wide range of biological specimens can be used, including body fluids (such as serum), secretions (like saliva), and excreta (such as urine or feces). Some samples can be tested directly, while others require pretreatment, especially fecal and certain secretion samples. Most ELISA tests use serum as the primary specimen. Plasma is similar to serum, except for the presence of fibrinogen and anticoagulants. To prepare plasma, anticoagulants must be used, while serum is obtained naturally through clotting. In most cases, serum is the preferred specimen for medical testing.
When collecting serum samples, care should be taken to avoid hemolysis, as red blood cells release substances with peroxidase activity, which can cause non-specific background staining in HRP-labeled assays. Serum should be tested fresh, and if stored, it should be kept at 4°C for up to five days or frozen for longer periods. Frozen serum should be gently mixed after thawing to prevent uneven distribution of proteins. Avoid shaking vigorously or creating air bubbles during mixing. If the serum shows signs of sedimentation or layering, it should be centrifuged or filtered before use.
Repeated freezing and thawing may reduce antibody titer, so it's advisable to divide serum samples into small aliquots for long-term storage. All steps should be performed under aseptic conditions. Additionally, reagents should be prepared according to the kit instructions, and distilled or deionized water should be of high quality for washing and other steps.
Before starting the ELISA test, make sure the reagents have reached room temperature. Unused parts of the kit should be returned to the refrigerator promptly. During the testing process, there are typically three addition steps: adding the sample, the enzyme conjugate, and the substrate. When adding the sample, it should be placed at the bottom of the well to avoid splashing or bubble formation. Use a micropipette for precise dosing, and replace the tip between each sample to prevent cross-contamination. For diluted samples, they can be mixed in a test tube before being added to the wells, and then shaken on a micro-vibrator for one minute to ensure even mixing.
For the enzyme conjugate and substrate solutions, a multi-channel pipette can be used to speed up the process. At Shanghai Jinma Experimental Equipment Co., Ltd., we specialize in providing high-quality ELISA kits, including wholesale options for both imported and domestic products. Our product range includes ELISA kits, antibodies, cell culture media, biological reagents, sera, standard reference materials, and more. With advanced technology and excellent customer service, we are committed to delivering top-tier solutions. Feel free to contact us for further discussion.