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The evaluation of ELISA reagents is divided into two aspects: one is the quality evaluation of the reagent itself, and the production can be produced after meeting certain requirements; the first is the evaluation of the effect in clinical application. Taking the hepatitis ELISA diagnostic reagent as an example, it is first necessary to pass the Chinese pharmaceutical biological product verification to obtain a production license. In addition to packaging, labels, instructions, etc., the performance of the reagents, such as specificity, sensitivity, precision and linearity, must be verified item by item. By testing a series of reference products, the results meet the requirements. . The clinical quality of the ELISA reagent is evaluated by using the reagent to observe the clinical application value. The Department of Clinical Examination conducted a work on the hepatitis B ELISA diagnostic reagent in this respect, and promoted the improvement of the reagent quality through quality evaluation.
I. Clinical quality evaluation points The reliability of the repair reagents is evaluated from the perspective of clinical application, based on its ability to distinguish between health and disease. In addition to packaging, labeling, instructions, etc., the functions of the reagents, such as specificity, agility, precision and linearity, need to be verified item by item. By testing a series of reference products, the results are in accordance with the requirements. qualified. The Central Inspection Center has carried out work on the hepatitis B ELISA diagnostic reagent in this respect, and promoted the improvement of reagent quality through quality evaluation. To perform this evaluation, it is first necessary to collect the relevant patient's serum and then determine it with the most reliable reagent for detecting the marker to determine whether it is positive or negative. It is still difficult to find a 100% reliable test, and any test will have false positives or false negatives. This group indicates that the test substance is a positive or negative serum composed of a "seed plate".
Second, the kit description
1. Avoid exposing the reagent to strong light during storage and incubation. All reagent bottle caps must be tightly closed to prevent evaporation and contamination, and the reagents should be protected from microbial contamination, as proteolytic enzyme interference can lead to erroneous results.
2. Carefully pipette the reagents and strictly follow the given incubation time and temperature. Please note that when taking specimens/standards, enzyme conjugates or substrates, the time interval between the first hole and the last hole loading will be too large, resulting in a different "pre-incubation" time, thus clearly Affect the accuracy and repeatability of the measured values. Moreover, insufficient washing will affect the test results.
3. Kit storage: Some reagents are stored at -20 °C, and some reagents are stored at 2-8 °C, which is subject to the label on the label.
4. The concentrated washing liquid will be salted out, and it can be heated and dissolved in the water bath when diluted.
5. The opening of the enzyme-linked plate well may contain a little water-like substance. This is a normal phenomenon and will not have any effect on the experimental results.
6. There may be inconsistencies in the Chinese and English manuals. Please refer to the English manual.
7. All samples should be managed and the samples and test equipment processed in accordance with the prescribed procedures.
8. Validity: 6 months 
Third, the elisa kit requires sample processing and requirements:
1. Serum: The blood is naturally coagulated at room temperature for 10-20 minutes and centrifuged for about 20 minutes (2000-3000 rpm). The supernatant is carefully collected, and if precipitation occurs during storage, it should be centrifuged again. Avoid any cellular irritation during the procedure. Use tubes without pyrogens and endotoxins.
2. Plasma: EDTA or sodium citrate should be selected as an anticoagulant according to the requirements of the specimen. After mixing for 10-20 minutes, the particles are removed by centrifugation for about 20 minutes (2000-3000 rpm). The supernatant is carefully collected, and if a precipitate forms during storage, it should be centrifuged again.
3. Urine: Collect with a sterile tube and centrifuge for about 20 minutes (2000-3000 rpm). The supernatant is carefully collected, and if a precipitate forms during storage, it should be centrifuged again. The chest and ascites and cerebrospinal fluid are referred to.
4. Cell culture supernatant: When detecting secreted components, collect them in a sterile tube. Centrifuge for about 20 minutes (2000-3000 rpm). Collect the supernatant carefully. When the components in the cells were detected, the cell suspension was diluted with PBS (pH 7.2-7.4), and the cell concentration reached about 1 million/ml. By repeated freezing and thawing, the cells are destroyed and the intracellular components are released. Centrifuge for about 20 minutes (2000-3000 rpm). Collect the supernatant carefully. If a precipitate forms during storage, it should be centrifuged again.
5. Tissue specimen: After cutting the specimen, weigh the weight. Add a certain amount of PBS, pH 7.4. It was quickly frozen and stored in liquid nitrogen for use. The specimen still maintains a temperature of 2-8 ° C after melting. A certain amount of PBS (pH 7.4) was added, and the specimen was homogenized by hand or homogenizer. Centrifuge for about 20 minutes (2000-3000 rpm). Collect the supernatant carefully. One part of the package is to be tested, and the rest is frozen for use.
6. The specimens should be extracted as soon as possible after collection, and the extraction should be carried out according to the relevant literature. The experiment should be carried out as soon as possible after extraction. If the test cannot be performed immediately, the specimen can be stored at -20 °C, but repeated freezing and thawing should be avoided.
7. Samples containing NaN3 cannot be detected because NaN3 inhibits horseradish peroxidase (HRP) activity.
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