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Adiponectin content detection kit and preparation method thereof

A detection kit and technology for adiponectin, which is applied in the field of adiponectin content detection kit, human body adiponectin content detection kit and its preparation, can solve the problem of batch detection of samples, low sensitivity, and kit Poor stability and other issues, to achieve the effect of large-scale promotion and application, simple detection operation, excellent sensitivity and specificity

Active Publication Date: 2016-12-21
WUHAN LIFE ORIGIN BIOTECH LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the use of radioactive raw materials, radioimmunoassay has serious environmental pollution problems, and the staff must take strict protective measures during the operation; enzyme-linked immunoassay has high detection accuracy, but the detection process is cumbersome and time-consuming Long, and samples need to be detected in batches, not suitable for timely detection; latex immunoturbidimetric method is also a common method for determining the concentration of adiponectin in human plasma or urine, it is based on common immunoturbidimetric method, but it overcomes the The defect of low sensitivity of ordinary immunoturbidimetric method, but also overcomes the defects of RIA and ELISA methods, has the advantages of simple operation, short detection time, high sensitivity, wide linear range, no pollution, wide application range, etc.
However, the existing adiponectin latex immunoturbidimetric assay kits have problems such as narrow linear range, poor anti-interference ability, and poor stability of the kit, which need to be improved.

Method used

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  • Adiponectin content detection kit and preparation method thereof
  • Adiponectin content detection kit and preparation method thereof
  • Adiponectin content detection kit and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] Preparation of Preliminary Example 1 Latex Particles Coated with Antibody to Adiponectin

[0049] (1) Latex washing: Take 1mL of latex solution with a concentration of 10% and a particle size of 80nm (purchased from PolyMicrospheres) in a 50mL centrifuge tube, then add 9mL of PBS buffer solution with pH7.4 into the centrifuge tube, shake and mix evenly, Then add 5 μL Tween-20 to the centrifuge tube, shake and mix evenly, place the centrifuge tube in a centrifuge at 25,000 rpm, centrifuge for 30 min, and discard the supernatant;

[0050] (2) Activation of latex: Reconstitute the latex precipitate in step (1) with 9 mL of PBS buffer at pH 7.4, and oscillate ultrasonically for 60 seconds to disperse the latex particles evenly. Accurately weigh 50 mg of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC) and 50 mg of N-hydroxysulfosuccinimide (sulfo-NHS) with 1 mL of PBS pH 7.4 Dissolve the buffer solution, add this solution into the reconstituted latex solution, shake and...

Embodiment 2

[0055] Preliminary Example 2 Preparation of latex particles coated with adiponectin antibody

[0056] (1) Latex washing: Take 1mL of latex solution with a concentration of 10% and a particle size of 200nm (purchased from PolyMicrospheres) in a 50mL centrifuge tube, then add 9mL of PBS buffer solution with pH6.5 into the centrifuge tube, shake and mix evenly, Then add 8 μL Tween-20 to the centrifuge tube, shake and mix evenly, place the centrifuge tube in a centrifuge at 25,000 rpm, centrifuge for 30 min, and discard the supernatant;

[0057] (2) Activation of latex: Reconstitute the latex precipitate in step (1) with 9 mL of PBS buffer at pH 6.5, and oscillate ultrasonically for 60 seconds to disperse the latex particles evenly. Accurately weigh 40mg of EDC and 40mg of NHS, dissolve it with 1mL of pH6.5 PBS buffer, add this solution into the reconstituted latex solution, oscillate and mix evenly, and place on a constant temperature shaker at 37 degrees, 200 rpm, and react 1h;...

Embodiment 3

[0062] Preliminary Example 3 Preparation of latex particles coated with adiponectin antibody

[0063] (1) Latex washing: Take 1mL of latex solution with a concentration of 10% and a particle size of 130nm (purchased from PolyMicrospheres) in a 50mL centrifuge tube, then add 9mL of PBS buffer solution with pH 7.0 to the centrifuge tube, shake and mix evenly, Then add 4 μL Tween-20 to the centrifuge tube, shake and mix evenly, place the centrifuge tube in a centrifuge at 25,000 rpm, centrifuge for 30 min, and discard the supernatant;

[0064] (2) Activation of latex: Reconstitute the latex precipitate prepared in step (1) with 9 mL of pH 7.0 PBS buffer, and oscillate ultrasonically for 60 seconds to disperse the latex particles evenly. Accurately weigh 55 mg of EDC and 55 mg of NHS, dissolve it with 1 mL of PBS buffer at pH 7.0, add this solution into the reconstituted latex solution, oscillate and mix evenly, and place on a constant temperature shaker at 37 degrees, 200 rpm, an...

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Abstract

The invention discloses an adiponectin content detection kit and a preparation method thereof. The adiponectin content detection kit comprises an independent reagent I and an independent reagent II. The reagent I comprises a bio-buffer, a surfactant, a coagulation accelerator, an antiseptic, a protective agent, a chelating agent and water. The reagent II comprises adiponectin antibody-coated latex particles, a bio-buffer, a chelating agent, a surfactant, an antiseptic, a suspending assistant, a blocking agent, a stabilizing agent, a protective agent and water. The adiponectin antibody-coated latex particles used in the adiponectin content detection kit have sizes of 80-200nm, and in the particle size range, detection sensitivity and a linearity range are obviously superior to those of the latex particles having other sizes. The sensitized latex particles are prepared by a chemical crosslinking method and the adiponectin antibody tightly bonds with the latex particle and difficultly fall off from the latex particle so that antibody bonding stability is improved and thus kit stability is improved.

Description

technical field [0001] The present invention relates to adiponectin content detection kit, in particular to a detection kit for adiponectin content in a human body and a preparation method thereof. The present invention further relates to a method for using the detection kit in detecting adiponectin content in a human body, The invention belongs to the detection field of adiponectin content. Background technique [0002] Adipose tissue is mainly composed of a large number of adipocytes aggregated into agglomerates, and adiponectin is an endogenous bioactive polypeptide or protein secreted by adipocytes. The concentration of adiponectin in normal human serum ranges from 3 to 30 μg / mL, accounting for 0.01% of the total plasma protein, and does not change with diet or circadian rhythm. The content of adiponectin in female serum is greater than that in males. With the continuous deepening of research on adiponectin, it has been found that the concentration of adiponectin in the...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/68G01N33/531
CPCG01N33/531G01N33/544G01N33/68
Inventor 华权高许可沈鹤霄黄爱舒芹鄢宝伍卫姣
Owner WUHAN LIFE ORIGIN BIOTECH LTD
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