Cytometric bead method for detecting blood platelet autoantibodies of immune thrombocytopenia

A technique for thrombocytopenia and autoantibodies, applied in measurement devices, instruments, disease diagnosis, etc., can solve the problems of low sensitivity, limited application of platelet antibodies, and high specificity, and achieve high sensitivity, convenient operation, and mature technology. Effect

Inactive Publication Date: 2017-12-01
冀学斌
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Problems solved by technology

These methods have high specificity and low sensitivity, which limits their application in clinical research of platelet antibodies

Method used

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Embodiment Construction

[0012] Detection steps of the present invention are as follows:

[0013] 1. Microsphere coating: The polystyrene microspheres are coated by physical adsorption method, and the Fab segment remains active by utilizing the hydrophobic effect of the Fc segment of the antibody to adsorb on the microspheres. Resuspend the microspheres before coating, add the microspheres to the mouse anti-human platelet membrane glycoprotein monoclonal antibody diluted in the coating buffer (make the ratio of antibody to microspheres 70ug antibody / ml microspheres), and vortex at room temperature for 2 Hours (or overnight at 4°C with vortex shaking), washed three times with 0.01M PBS / TWEEN, then blocked the microspheres with 5% bovine serum albumin shaking for 2 hours, washed three times with 0.01M PBS / TWEEN, and washed in an isotonic medium containing 0.2% sodium azide Resuspend the microspheres in saline solution to a final concentration of 0.25% microspheres. Store at 4°C. On days 1, 2, 5, 10, 3...

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Abstract

The invention belongs to blood platelet clinical test and research and relates to a cytometric bead method for detecting blood platelet autoantibodies of immune thrombocytopenia. The cytometric bead method for detecting blood platelet autoantibodies of immune thrombocytopenia is high in sensitivity and specificity. The method comprises encapsulating beads through anti-human platelet membrane glycoprotein monoclonal antibodies, incubating platelet lysate and the encapsulated beads, and then adding in phycoerythrin marked goat antihuman immune globulin polyclonal antibodies to perform flow cytometry. If autoantibodies exist on the surface of platelets, a bead-platelet membrane glycoprotein monoclonal antibody-platelet membrane glycoprotein autoantibody-phycoerythrin marked goat antihuman immune globulin polyclonal antibody composite structure can be formed and embodied as increase of fluorescence intensity of detecting beads. The cytometric bead method for detecting blood platelet autoantibodies of immune thrombocytopenia is convenient to operate, mature in technology, high in sensitivity of blood platelet autoantibody detection and conductive to fundamental experimental research on platelet antibodies.

Description

Technical field: [0001] The invention belongs to a novel test method with high sensitivity and specificity for detecting platelet-specific antibodies in platelet clinical test research. Background technique: [0002] Thrombocytopenia is a common disease in clinical hematology. According to different pathogenesis, it can be divided into immune thrombocytopenia and non-immune thrombocytopenia. How to distinguish between the two requires a platelet autoantibody test method with high specificity and sensitivity. [0003] According to the time when the platelet antibody detection method appears in the laboratory, it can be divided into phase I, phase II and phase III. Phase I examination appeared from the 1950s to the 1970s. It mainly detects the effect of patient serum or plasma on normal platelet function in vitro. The test methods used are all indirect methods with low sensitivity and specificity. Phase II assays were developed in the 1970s to detect platelet-associated ant...

Claims

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Application Information

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IPC IPC(8): G01N33/543
CPCG01N33/54313G01N2800/222
Inventor 冀学斌李秋环张丽萍
Owner 冀学斌
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