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Kit for specifically detecting creatine kinase isoenzyme

A technology of creatine kinase and isoenzyme, applied in the field of creatine kinase detection, can solve the problems of low detection linear range and low detection specificity of creatine kinase isoenzyme detection kit, and achieve high linearity and resistance to HOOK effect. , Improve the detection linear range, the effect of high sensitivity

Active Publication Date: 2021-07-30
BEIJING DIAGREAT BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Aiming at the problem of low detection specificity and low detection linear range of related creatine kinase isoenzyme detection kits, this application provides a specific detection kit for creatine kinase isoenzyme

Method used

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  • Kit for specifically detecting creatine kinase isoenzyme
  • Kit for specifically detecting creatine kinase isoenzyme
  • Kit for specifically detecting creatine kinase isoenzyme

Examples

Experimental program
Comparison scheme
Effect test

preparation example 1

[0040] Preparation of Reagent RM

[0041] (1) Use a pipette gun to measure 1.5mL magnetic microspheres (10% solid content) into the corresponding centrifuge tube, magnetically absorb it on the magnetic stand for 3min, discard the supernatant, add 5mL coating buffer, and vortex to mix 10 seconds, then magnetic suction for 3 minutes, discard the supernatant, add 4mL coating buffer; the coating buffer is 50mmol / L phosphate buffer;

[0042] (2) Add 2mg of CKMM coating antibody, vortex for 10s, place in a shaker and mix in a 37°C drying oven for 4h (shaker parameters: 1000rpm);

[0043] (3) Add 1mL of blocking solution, place in a shaker and mix in a 37°C drying oven for 4h (shaker parameters: 1000rpm);

[0044] (4) Magnetic suction for 3 minutes, remove the supernatant, use 10mL magnetic bead preservation buffer and invert repeatedly 20 times, mix well;

[0045] The components of the magnetic bead preservation buffer include 20mmol / L Tris buffer, 0.1% BSA, 0.1% Tween20, 0.1% polye...

preparation example 2

[0048] Preparation of Reagent RM

[0049] (1) Use a pipette gun to measure 1.5mL magnetic microspheres (10% solid content) into the corresponding centrifuge tube, magnetically absorb it on the magnetic stand for 3min, discard the supernatant, add 5mL coating buffer, and vortex to mix 10 seconds, then magnetic suction for 3 minutes, discard the supernatant, add 4mL coating buffer; the coating buffer is 50mmol / L phosphate buffer;

[0050] (2) Add 2mg of CKMM coating antibody, vortex for 10s, place in a shaker and mix in a 37°C drying oven for 4h (shaker parameters: 1000rpm);

[0051] (3) Add 1mL of blocking solution, place in a shaker and mix in a 37°C drying oven for 4h (shaker parameters: 1000rpm);

[0052] (4) Magnetic suction for 3 minutes, remove the supernatant, use 10mL magnetic bead preservation buffer and invert repeatedly 20 times, mix well;

[0053] The components of the magnetic bead preservation buffer include 20mmol / L Tris buffer, 0.1% BSA, 0.1% Tween20, 0.1% pol...

preparation example 3

[0056] Preparation of Reagent R1

[0057] (1) Take 1 mg of CKMM-labeled antibody, add 1 mg of EDC (EDC is prepared with water to 10 mg / mL, ready-to-use), and stand at room temperature for 20 minutes;

[0058] (2) Take 2mg of alkaline phosphatase and add appropriate amount of purified water to dilute to 1mg / mL; add purified water (mL)=2-(2 / C), add alkaline phosphatase volume (mL)=2 / C, wherein, C (mg / mL) is the concentration of alkaline phosphatase;

[0059] (3) Mix the mixture obtained in step (1) and the mixture obtained in step (2), and leave to react at room temperature for 60 minutes;

[0060] (4) Place the mixture obtained after standing in step (3) into a dialysis bag with a pore size of 7K, and place it in 500 mL of enzyme marker storage buffer, place it at 2-8°C for 4 hours for dialysis, and repeat this step twice;

[0061] The components of the enzyme marker preservation buffer include 25mmol / L Tris buffer, 0.1% BSA, 0.1% Tween20, 0.1% polyethylene glycol 6000, 0.01%...

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Abstract

The invention discloses a kit for specifically detecting creatine kinase isoenzyme and belongs to the technical field of creatine kinase detection. The kit comprises a reagent R0, a reagent RM and a reagent R1, wherein the reagent R0 comprises a buffer solution, sodium chloride, a protective agent, a preservative and a creatine kinase isoenzyme B subunit antibody; the reagent RM comprises a buffer solution, magnetic microspheres coated with a creatine kinase MM isoenzyme coating antibody, a protective agent, a surfactant, a stabilizer and a preservative; the reagent R1 comprises a buffer solution, a creatine kinase MM isoenzyme labeled antibody coated with alkaline phosphatase, a protective agent, a surfactant, a stabilizer and a preservative. The kit is high in sensitivity and good in specificity, has a relatively large linear range and HOOK resistance, can detect CKMM in a dry filter blood sheet, serum or urine sample, and is relatively wide in application range.

Description

technical field [0001] The application relates to the technical field of creatine kinase detection, in particular to a kit for specifically detecting creatine kinase isozymes. Background technique [0002] Creatine Kinase (Creatine Kinase, CK) is an important kinase that is directly related to intracellular energy operation, muscle contraction, and ATP regeneration. It is mainly distributed in skeletal muscle and cardiac muscle, followed by brain tissue. For the diagnosis of skeletal muscle disease and myocardial disease. There are four isozymes of creatine kinase, CKMM is the muscle isozyme, mainly distributed in skeletal muscle; CKMB is the heart isozyme, mainly distributed in the myocardium; CKBB is the brain isozyme, mainly distributed in Brain tissue; CKMiMi is a mitochondrial isoenzyme, mainly distributed in the mitochondria of cardiac muscle and skeletal muscle. Among them, CKMM has great diagnostic value for muscle injury and neonatal Duchenne muscular dystrophy. ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/58G01N33/573G01N33/543
CPCG01N33/581G01N33/573G01N33/54326G01N2333/9123G01N2800/2892
Inventor 吴鸣月常缘荣王艳新周裕军
Owner BEIJING DIAGREAT BIOTECH CO LTD
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