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Kit for detecting short-chain fatty acid and application thereof

A technology of short-chain fatty acids and kits, which can be used in measuring devices, instruments, scientific instruments, etc., and can solve the problems of low sensitivity and instability of kits

Pending Publication Date: 2020-03-24
山东莱博生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The relevant ELISA kits reported in the literature are all horseradish peroxidase (HRP) markers, which are relatively unstable and the sensitivity of the kits is not high
There is no report on the enzyme-linked immunoassay kit for detecting short-chain fatty acids with alkaline phosphatase (ALP) as a marker, simple operation, stable marker, and high sensitivity

Method used

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  • Kit for detecting short-chain fatty acid and application thereof
  • Kit for detecting short-chain fatty acid and application thereof
  • Kit for detecting short-chain fatty acid and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0069] The test kit for detecting short-chain fatty acids of the present invention includes an enzyme label plate, a gradient solution of short-chain fatty acid standard products, a sample diluent, and an alkaline phosphatase (ALP)-labeled detection-resistant short-chain fatty acid conjugate in the box. Antibody, 20× concentrated washing solution, chromogenic reagent, stop solution and plate sealing film, sealed bag and instructions;

[0070] in:

[0071] The microtiter plate is a transparent polystyrene 96- or 48-well microtiter plate, and each well is coated with an anti-short-chain fatty acid monoclonal antibody Mab1 at a concentration of 5 μg / mL;

[0072] Described short-chain fatty acid standard substance gradient solution has the solution of 6 concentrations, makes as follows: by acetic acid: propionic acid: butyric acid: valeric acid: hexanoic acid=60mg / ml:25mg / ml:14mg / ml:0.5 mg / ml:0.5mg / ml ratio, prepare the short-chain fatty acid standard substance in simulated serum...

Embodiment 2

[0080] The preparation method of the kit for detecting short-chain fatty acids of the present invention, the steps are:

[0081] 1) Preparation of enzyme plate:

[0082] 1.1) Preparation of coating solution: prepare 0.1mol / L sodium carbonate-sodium bicarbonate buffer solution, and adjust the pH value to 9.0; weigh 1 g of α-cyclodextrin and add it to 1L sodium carbonate-sodium bicarbonate buffer solution, Prepare coating buffer;

[0083] 1.2) prepare washing liquid, the formula of washing liquid and the preparation method are:

[0084]

[0085] Sterilize by filtration and store at 4°C;

[0086] 1.3) Prepare blocking solution, the formula and preparation method of blocking solution are:

[0087]

[0088] Sterilize by filtration and store at 4°C;

[0089] 1.4) Dilute the anti-short-chain fatty acid monoclonal antibody Mab1 with the coating solution to a working concentration of 5 μg / mL, mix well, and let stand for 15 minutes;

[0090] 1.5) Take the marked ELISA plate, ...

Embodiment 3

[0107] The application of the kit for detecting short-chain fatty acids of the present invention in detecting short-chain fatty acids.

[0108] The specific steps for detecting short-chain fatty acids are:

[0109] 1) Equilibration: equilibrate the sample to be tested and the reagents in the detection kit at 18-25°C for 30±5 minutes;

[0110] 2) Adding samples of standard substance: set the standard substance well and the sample well, and add 50 μL of standard substance of different concentrations to the standard substance well according to the concentration gradient;

[0111] 3) Adding samples: set blank wells and sample wells to be tested on the enzyme-labeled plate; add 40 μL of sample diluent to the sample wells to be tested, and then add 10 μL of sample to be tested to make the final dilution of the sample 5 times; When sampling, add the sample to the bottom of the well of the plate, try not to touch the wall of the well, shake gently to mix; do not add sample and enzyme...

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Abstract

The invention discloses a kit for detecting short-chain fatty acid. The kit comprises a kit body, an elisa plate arranged in the kit body, a short-chain fatty acid standard substance gradient solution, a sample diluent, an alkaline phosphatase labeled antibody for detecting an anti-short-chain fatty acid conjugate, a 20* concentrated washing solution, a color developing agent, a stop solution, a plate sealing film, a sealing bag and an instruction book, wherein the elisa plate is a transparent polystyrene 96 or 48-hole elisa plate, and each hole of the elisa plate is coated with an anti-short-chain fatty acid monoclonal antibody Mab1 with the concentration of 5mu g / mL. The short-chain fatty acid standard substance gradient solution has six concentrations, separately 20 ng / mL, 10 ng / mL, 5 ng / mL, 2.5 ng / mL, 1.25 ng / mL and 0 ng / mL. The detection kit provided by the invention provides an enzyme-labeled secondary antibody labeled by alkaline phosphatase (ALP), so that the stability and sensitivity of the kit are remarkably improved.

Description

technical field [0001] The invention relates to a kit for detecting metabolites of gastrointestinal flora, in particular to an enzyme-linked immunoassay kit for detecting short-chain fatty acids (SCFAs) with alkaline phosphatase (ALP) as a marker and application thereof. Background technique [0002] Short-chain fatty acids (SCFAs) are end products of dietary fiber and some amino acids and proteins fermented by probiotics, mainly intestinal anaerobic probiotics, mainly including acetic acid, propionic acid, isobutyric acid, butyric acid , Isovaleric acid, valeric acid and caproic acid, etc. Short-chain fatty acids can not only provide energy for the cells of the body, but also have the functions of inhibiting pathogenic bacteria, inhibiting the inflammatory response of intestinal epithelial cells and bone marrow cells, and can regulate the body's metabolic balance, reducing inflammatory bowel disease, obesity and type II diabetes and other disease incidence. [0003] At pr...

Claims

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Application Information

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IPC IPC(8): G01N33/577G01N33/535
CPCG01N33/577G01N33/535
Inventor 陈振朱之炜欧兰香郎伟超王佳颖
Owner 山东莱博生物科技有限公司
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