Sensor detection kit based on 96-well plate immobilized molecular motor and method for in-vitro detection of P24 antigen

A technology for detection kits and molecular motors, applied in instruments, measuring devices, scientific instruments, etc., can solve problems such as high repeatability, cumbersome and complicated operations, and large batch-to-batch differences, and achieve good uniformity, strong specificity, and detection The effect of high sensitivity

Pending Publication Date: 2022-07-29
ZHEJIANG UNIV OF TECH +1
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Problems solved by technology

[0003] Traditional molecular motor biosensor detection kits often immobilize molecular motors on the surface of magnetic beads to construct biosensors, but this reconstruction method relies on the biotin-streptavidin-biological system to connect magnetic beads and molecular motors, which is e...

Method used

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  • Sensor detection kit based on 96-well plate immobilized molecular motor and method for in-vitro detection of P24 antigen
  • Sensor detection kit based on 96-well plate immobilized molecular motor and method for in-vitro detection of P24 antigen
  • Sensor detection kit based on 96-well plate immobilized molecular motor and method for in-vitro detection of P24 antigen

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Embodiment

[0039] The preparation process of biotinylated subunit monoclonal antibody or biotinylated P24 antibody in this example is as follows: biotin and P24 antibody or subunit monoclonal antibody are mixed in carbonate buffer (0.1mol / mol / min) in a mass ratio of 10:1. L, pH 9.2) mixed for 1 h, and then collected by dialysis.

[0040] 1. Preparation of biotinylated molecular motors:

[0041] Take 10 mg of 16:0 Biotinyl PE, dissolve it in 1 ml of chloroform, prepare a 10 mg / ml Biotinyl PE solution, and take 5 ml of F0F1-ATPase molecular motor solution (the molecular motor concentration is expressed as synthetic activity, and the synthetic activity is 623789 ) were sonicated for 10 min at an ultrasonic frequency of 200 W, and the above Biotinyl PE solution was added dropwise during the ultrasonic process to prepare a biotinylated molecular motor solution and stored at 4 °C for later use.

[0042] 2. Test plate preparation

[0043]Add 100 μg / ml streptavidin solution to a 96-well white ...

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Abstract

The invention discloses a sensor detection kit based on a 96-well plate immobilized molecular motor and a method for P24 antigen in vitro detection.The kit is composed of a molecular motor sensor detection plate, and the detection plate is formed by immobilizing an F0F1-ATP enzyme molecular motor on a 96-well ELISA plate; meanwhile, a detection probe (P24 antibody) is combined to form a molecular motor biosensor for capturing a target. The detection method mainly comprises the following steps: (1) placing a sample in the detection plate, and capturing a target object by the sensor; (2) adding an ATP (adenosine triphosphate) synthesis solution containing ADP (adenosine triphosphate), and reacting And (3) adding an ATP detection reagent, quickly and uniformly mixing, and starting a chemical light emission detector to measure the luminous intensity. According to the present invention, the stability and the uniformity of the 96-well plate immobilized molecular motor, the mutual conversion characteristic of the kinetic energy and the chemical energy of the molecular motor, and the high sensitivity and the rapid performance of the chemiluminescence are utilized to detect the antigen, and the porosity of the 96-well plate enables the rapid and stable detection to be achieved while the high throughput screening requirement can be met.

Description

technical field [0001] The invention belongs to the field of in vitro diagnostic reagents, in particular to a sensor detection kit based on a 96-well plate immobilized molecular motor, and a method for in vitro detection of P24 antigen. Background technique [0002] Biomolecular motors are proteases that convert chemical energy into mechanical energy. Among them, ATP synthase, also known as F0F1-ATP synthase (F0F1-ATPase), is widely present in the mitochondrial inner membrane of animals and fungi, plant chloroplast thylakoid membranes and bacteria. The plasma membrane is the main site of ATP synthesis and is the smallest known rotary molecular motor. F0F1-ATP synthase can not only synthesize ATP using the transmembrane proton gradient and drive the rotor to rotate clockwise, but also hydrolyze ATP to transport protons and drive the rotor to rotate counterclockwise. Due to the special motion and rotation characteristics of F0F1-ATPase, it can be prepared as a molecular motor...

Claims

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Application Information

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IPC IPC(8): G01N33/543G01N33/533
CPCG01N33/54326G01N33/54346G01N33/54366G01N33/533
Inventor 杨根生应三军高颖洪伟勇楼邦
Owner ZHEJIANG UNIV OF TECH
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