Competitive homogeneous chemiluminescence assay kit and application thereof
A homogeneous chemiluminescence and chemiluminescence technology, which is applied in the direction of measuring devices, scientific instruments, instruments, etc., can solve the problems of not being able to effectively take into account functional sensitivity and analysis range, and achieve broadened detection range, high precision and accuracy, and guaranteed The effect of functional sensitivity
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Embodiment 1
[0073] Embodiment 1: for the preparation of the kit of the present invention that the analyte is estradiol (E2)
[0074] (1) Preparation process of monoclonal antibody-coupled receptor microsphere solution (reagent 1)
[0075] Receptor microspheres: The surface of the microspheres contains aldehyde groups (-CHO), which are connected to antibody molecules through aldehyde groups. Contains luminescent compounds (derivatives of dimethylthiophene) and chelates of lanthanide (Eu) compounds.
[0076] Biological material: a high-affinity monoclonal antibody (ie, HA-McAb) that specifically binds E2 and a low-affinity monoclonal antibody that specifically binds E2 (ie, LA-McAb).
[0077] Preparation process: Preparation process:
[0078] 1) Monoclonal antibody with high affinity to E2 (ie, HA-McAb) was dialyzed overnight with carbonate buffer, mixed with acceptor microspheres (FG), and the mass ratio of antibody to microspheres was 1 :400, coated for 2 hours, added blocking solution...
Embodiment 2
[0090] Using the method in Example 1, the conditions such as antibody type, coupling mass ratio, and competing antigen type were changed, and the same batch of samples was detected by the LiCA500 automatic photochemiluminescence analysis system, and the homogeneous chemiluminescence signal was automatically completed and output. The detection range and detection limit of the test results.
[0091] The detection process using the kit prepared in Example 1 is fully automated by the LiCA500 automatic light-activated chemiluminescence analysis system and the detection results are output. The specific steps are:
[0092] a. Add 10 μl of sample, calibrator or quality control to the reaction well;
[0093] b. Add 25 μl release agent, 25 μl reagent 1 and 25 μl reagent 2 to the reaction well in sequence;
[0094] c. Incubate at 37°C for 15 minutes;
[0095] d. Add 175 μl of LiCA universal solution (donor microsphere solution combined with avidin);
[0096] e. Incubate at 37°C for 15...
Embodiment 3
[0103] The precision of the kit used in Test No. 7 in Example 2, in which Bio-E2 was used as a competing antigen, was examined.
[0104] Significance of precision: Precision is an important indicator to measure the intra-assay and inter-assay variation of the kit, and is an important basis for evaluating the effectiveness of the product to be marketed, usually including intra-assay precision and inter-assay precision.
[0105] Intra-batch precision evaluation method: use low (L), medium (M), and high (H) value samples to conduct independent analysis on a batch of products, repeat the measurement 10 times for each batch, and calculate the measurement results of 10 times average of and standard deviation (SD), according to the formula The coefficient of variation (CV) was calculated and the results are shown in Table 1.
[0106] Inter-batch precision evaluation method: Use samples with low (L), medium (M), and high (H) values to conduct independent analysis on 3 batches of...
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