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Method for detecting activated blood coagulation factor XI in human intravenous immunoglobulin

A technology for immunoglobulin and blood coagulation factors, which is applied in material excitation analysis, fluorescence/phosphorescence, etc., can solve the problems of inability to accurately measure the specific content of activated blood coagulation factors in samples, and inability to monitor changes in the thrombin generation process in real time

Active Publication Date: 2013-07-03
BLOOD TRASFUSION INST CHINESE ACAD OF MEDICAL SCI +1
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Problems solved by technology

At present, this method has the following shortcomings: ①It can only be qualitative, and cannot monitor the changes of the thrombin generation process in the sample in real time; ②The result can only be judged by the clotting time of the sample, and human factors are greatly affected, so it is impossible to accurately determine the specific content of activated coagulation factors in the sample. content

Method used

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Embodiment Construction

[0014] The method of the present invention will be further described in detail below in conjunction with specific examples.

[0015] 1. Preparation of FXIa-free PPP

[0016] Plasma was collected from healthy blood donors using venipuncture. Add corn trypsin inhibitor at a concentration of 40-150 μg / ml to sodium citrate anticoagulated plasma at a ratio of 1:9, mix well, and then centrifuge at 4°C at 1500 rpm for 30 min. Take the upper 2 / 3 plasma with a plastic syringe, centrifuge at 3500 rpm, 4°C for 30 minutes, take the upper 2 / 3 plasma, divide it into plastic tubes, 2ml each, and store at -40°C for later use;

[0017] 2. Establish a TGT-based FXIa detection method

[0018] (1) Optimizing the substrate concentration

[0019] The choice of thrombin-specific fluorescent substrate Boc-Val-Arg-AMC.HCL can stably and specifically reflect the changes in the TGT generation process. The thrombin-specific fluorescent substrate Boc-Val-Arg-AMC.HCL is insoluble in Hepes buffer, so it...

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Abstract

The invention discloses a method for detecting an activated blood coagulation factor XI in human intravenous immunoglobulin. The method comprises the steps as follows: (1) mixing an IVIG (intravenous immunoglobulin) sample with specially-preprocessed platelet-poor plasma (PPP); (2) adding a thrombin specific fluorogenic substrate with a phospholipid agent into a reaction system; (3) adding an initiation factor for starting a TGT (thrombin generation test), wherein the initiation factor comprises a calcium ion source and the phospholipid agent; (4) obtaining a thrombin generation curve providing parameters by the optimized reaction system; and (5) negatively correlating the peak time of thrombin (TTP) of the detected sample with the FXIa (activated blood coagulation factor XI) level in the sample. The FXIa level in the detected sample can be indirectly obtained by comparing the TTP of the detected sample with that of an FXIa standard sample. The method is suitable for detecting residual micro FXIa in human intravenous immunoglobulin (IVIG) products and is used for extracorporal evaluation of thrombosis risk of related products.

Description

technical field [0001] The invention belongs to the technical field of biomedicine, and relates to a detection method for activated coagulation factor XI (hereinafter referred to as FXIa) in human intravenous immunoglobulin (pH4, Human intravenous immunoglobulin, hereinafter referred to as IVIG). This method can accurately detect the specific content of FXIa in IVIG, and the sensitivity can reach 0.12 nmol / L. Background technique [0002] IVIG is the immunoglobulin component extracted from the plasma of healthy people immunized with hepatitis B vaccine by the low-temperature ethanol protein separation method, and refined through further processing and virus inactivation. IVIG has dual therapeutic effects of immune replacement and immune regulation, and is widely used clinically. In recent years, with the increase in the number of IVIG clinical use, the adverse reaction events after the product is infused into the human body are also gradually increasing, and the harm is ser...

Claims

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Application Information

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IPC IPC(8): G01N21/64
Inventor 马莉孙盼李长清袁靖陈云华刘欣晏杨刚
Owner BLOOD TRASFUSION INST CHINESE ACAD OF MEDICAL SCI
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