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Serum marker capable of evaluating cerebral hemorrhage risk before thrombolysis and application thereof

A marker and serum technology, applied in measurement devices, instruments, scientific instruments, etc., to maintain the natural conformation and activity, overcome surface tension and steric barriers, and facilitate adequate responses

Inactive Publication Date: 2015-12-30
THE SECOND PEOPLES HOSPITAL OF SHENZHEN
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, at present, Occludin protein has not been used in the detection of peripheral blood before thrombolysis (administration), so as to judge the risk of cerebral hemorrhage after thrombolysis, and to evaluate the risk of cerebral hemorrhage after thrombolysis

Method used

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  • Serum marker capable of evaluating cerebral hemorrhage risk before thrombolysis and application thereof
  • Serum marker capable of evaluating cerebral hemorrhage risk before thrombolysis and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0054] Serum samples were collected, and the occludin protein concentration in the serum was measured after the occurrence of cerebral ischemia with the basic level as the control. The results were as follows: figure 1 shown. in, figure 1 A shows that the basal level of Occludin protein in serum is low, and after 4 hours of middle cerebral artery embolism (MCAO), the level of Occludin protein fragments in peripheral blood increases significantly compared with the sham operation group; figure 1 In B, the 55kDa protein fragment was increased 2 hours after the middle cerebral artery occlusion (MCAO), and the 31kDa Occludin protein fragment was detected later, and it was not obvious until 4 hours after the middle cerebral artery occlusion (MCAO).

Embodiment 2

[0056] The method for preparing the test kit for detecting the Occludin protein in human serum is as follows:

[0057] Step 1: Preparation of Microsphere Storage Solution

[0058] (1) Activation of microspheres: after the polystyrene microspheres are uniformly dispersed, take 5.0×10 6 Add 100 μl washing buffer (PBS, 0.05% Tween) to the microspheres, shake at high speed for 15 seconds, suspend the microspheres with 80 μL activation buffer after washing, add 10 μL carbodiimide (EDC, 50 mg / ml), and quickly add 10 μL Sulfo-NHS (50mg / ml), shake at high speed for 30s, shake at room temperature for 20min in the dark, then wash twice with 500μL 0.01MpH7.4PBS, centrifuge at 12000r / min for 3min each time, discard the supernatant, and finally suspend the microspheres with 100μL0.01MpH7.4PBS .

[0059] (2) Cross-linking of microspheres: Add 100 μg of anti-Occludin protein monoclonal antibody to the activated microspheres, dilute to 500 μL with 0.01M pH7.4 PBS, wrap in tinfoil, shake at ...

Embodiment 3

[0080] 1. the test kit of embodiment 2, by 1 piece of 96 hole filter plate, 2 bottles of standard substance, 1 bottle of sample diluent, the polystyrene microsphere that the surface has been coupled with anti-GFAP antibody, 1 bottle of microsphere storage solution , 1 bottle of enzyme conjugate, 1 bottle of substrate A1 bottle, 1 bottle of substrate B1, 20× washing solution, 20 self-adhesive coverslips and 1 instruction manual.

[0081] 2. the application of the test kit of embodiment 2

[0082] (1) Dilution of the standard substance: Dilute each bottle to 1mL with the sample diluent before use. After standing for 10 minutes, mix it upside down repeatedly. mL, 12.5ng / mL, 3.12ng / mL, 0.78ng / mL, 0.05ng / mL, the sample diluent is 0ng / mL.

[0083] (2) Adding samples: Shake and mix the microspheres at a medium speed before use, add 10 μL microspheres containing about 50,000 Occludin protein monoclonal antibodies to each well, vacuum filter and wash twice with 100 μL washing liquid, an...

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Abstract

The invention relates to a serum marker capable of evaluating a cerebral hemorrhage risk before thrombolysis and application thereof and provides a kit used for detecting Occludin protein in the serum in the human body. When the kit comprising test strips is adopted, the kit has the advantages of being rapid, easy and convenient to use, accurate, low in cost, good in specificity and high in sensitivity, and the Occludin protein level result in peripheral blood before thrombolysis can be read within 5-15 min; when the kit comprising a 96-hole filter plate is adopted, the kit also has the advantage of being capable of rapidly testing the Occludin protein level in peripheral blood before thrombolysis. By adopting the kit for detecting the Occludin protein level in peripheral blood before thrombolysis (dosing), the risk that cerebral hemorrhage possibly occurs after thrombolysis can be effectively judged, a doctor is guided for medicine usage, and the great clinical significance is achieved.

Description

technical field [0001] The invention relates to the field of immunoassay and analysis, in particular to a kit and its application, in particular to a serum marker capable of assessing the risk of cerebral hemorrhage before thrombolysis and its application. Background technique [0002] Acute ischemic stroke is a common and frequently-occurring disease among middle-aged and elderly people, with high disability and mortality rates. Early thrombolysis with tissue plasminogen activator (tPA) is currently the only ischemic stroke approved by the US FDA. stroke therapy. However, the risk of intracerebral hemorrhage after thrombolysis has always been considered by clinicians. Once severe cerebral hemorrhage occurs, the mortality rate of patients is as high as 80%. Therefore, how to reduce the occurrence of cerebral hemorrhage and increase the safety of tPA thrombolytic therapy has been an important clinical issue that has plagued the clinic for a long time. [0003] At present, t...

Claims

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Application Information

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IPC IPC(8): G01N33/535
CPCG01N33/535
Inventor 刘文兰徐迹李维平
Owner THE SECOND PEOPLES HOSPITAL OF SHENZHEN
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