Method and kit for cross-over lymphatic toxicity experiment

Abstract

The invention discloses a method for a cross-over lymphatic toxicity experiment. The method comprises the following steps: (1) taking the venous anticoagulant whole blood of a transplant donor and theserum of a transplant recipient, and separating donor lymphocytes from the venous anticoagulant whole blood of the transplant donor; (2) taking a donor lymphocyte fluid, carrying out staining and marking, and adjusting the cell concentration; (3) taking and dissolving lyophilized complement; (4) taking five reaction tubes, sequentially adding two or three of the donor lymphocytes, normal saline,positive serum and the recipient serum, carrying out uniform mixing, standing the obtained mixture, adding a fifth reagent rabbit complement, uniformly mixing the mixture and the rabbit complement, and standing the new mixture; and (5) staining the new mixture, and loading the obtained sample. The invention also discloses a detection kit for the cross-over lymphatic toxicity experiment. The methodand the kit have the advantages of low toxicity, low cost, high detection accuracy and excellent clinical application prospect.

Description

technical field [0001] The invention relates to a method and kit for cross-lymphatic toxicity experiment. Background technique [0002] The cross-lymphocytotoxicity test is the complement-dependent cytotoxicity test (CDC), which is used to determine whether the recipient has pre-existing antibodies against the donor before allogeneic organ transplantation. Pre-existing antibodies in the recipient may be produced due to multiple blood transfusions, pregnancy, secondary transplantation and other reasons. If there are cytotoxic antibodies in the recipient, it may cause hyperacute rejection after transplantation, resulting in transplant failure. Therefore, it is necessary to check whether there are anti-donor lymphocyte antibodies in the recipient's serum before transplantation to prevent hyperacute rejection. experiment. [0003] In 1969, Patel and Terasaki invented the technique of complement-dependent cytotoxicity test for the screening of acute and hyperacute rejection bef...

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Problems solved by technology

[0005] Fluorescence staining detection method is usually through the microplate method, the material used is Terasaki microplate, which needs to be observed and judged by an inverted fluorescence microscope, and there are the following three defects: 1. Terasaki microplate is a special reaction plate, and the space of the reaction system is limited, which is not conducive to The full mixing and reaction of each experimental component also easily causes errors in counting and typing due to cell overlap during counting; 2. Inverted fluorescence microscopes are expensive, and hospitals that carry out this project need to spend hundreds of thousands of dollars to purchase , which limits the development of this experimental method; 3. The fluorescent dye solution used contains ethidium bromide, which is carcinogenic, and prolonged use may cause cancer

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