Kit used for detecting beta-lactamase in dairy and detection method thereof

A technology for lactamase and detection reagents, which is applied in the field of β-lactamase kits and its detection, can solve the problems of high detection cost, high cost, and unclear interpretation of results, etc.

Inactive Publication Date: 2017-04-05
北京华益精点生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] For β-lactamase, the principles of existing detection methods are mostly based on ELISA, physical and chemical methods, microbial methods and instrumental detection. The detection sensitivity of ELISA depends on the purity and specificity of receptors and ligands and the cost. High; the results of physical and chemical methods are subject to the operation of personnel to a certain extent; instrumental methods require specific equipment and the detection cost is expensive; microbial methods have better overall results in the detection of β-lactamases, such as the paper recommended by NCCLS in the United States The slice diffusion method, but the interpretation of the results of the microbiological method is based on the observation of the inhibition zone, which is not very clear in the interpretation of the results, and there will be interpretation errors due to human factors

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0030] Embodiment 1: Preparation of β-lactamase detection kit

[0031] A β-lactamase detection kit, which consists of a detection body and a detection reagent. The detection body is a coated 96-well plate containing detection bacteria, acid-base indicators, and nutrients required for bacterial growth. , the detection reagent is an organic solution containing β-lactam antibiotics.

[0032] The detection bacterium is a suspension of Bacillus stearothermophilus;

[0033] The acid-base indicator is bromocresol purple;

[0034] The detection reagent is piperacillin dissolved in acetonitrile with a concentration of 10 g / mL and stored at -20°C.

[0035] 1. Preparation of Assay Subjects

[0036] Preparation of spore suspension

[0037] a. Plate method:

[0038] 5) Take a colony from the activated Bacillus stearothermophilus colony and inoculate it into a test tube containing a sterile liquid medium, and shake the bacteria at 56-65°C for about 24 hours;

[0039] 6) On the next da...

Embodiment 2

[0057] Embodiment 2: sample detection

[0058] Take 1 microliter of detection reagent and add it to 1 milliliter of the milk sample to be tested, mix evenly and pre-incubate at 40°C for 1 hour; Seal and incubate again at 64°C for about 3 hours. If the milk sample to be tested contains β-lactamase and the content is above the detection limit, the active ingredients in the detection reagent will be degraded after the pretreatment stage, the growth of microorganisms will not be inhibited, and the bacteria will produce acid to make the indicator in the micropores On the contrary, if the milk sample to be tested does not contain β-lactamase or the content is below the detection limit, the milk sample as a whole still has antibacterial properties, microorganisms cannot grow and produce acid, and the indicator in the micropores Keep the original purple or blue-purple.

Embodiment 3

[0059] Embodiment 3: the test of kit minimum detection line

[0060] Take 1ml of 4.46 million IU / ml β-lactamase (bought from the market); dilute with a negative milk sample to obtain a milk sample containing 4IU / ml; operate according to the operation steps of the kit; the test result β-lactamase Amidase was positive.

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PUM

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Abstract

The invention discloses a kit used for detecting beta-lactamase in dairy and a detection method thereof. The kit is composed of two parts of a detection main body and a detection reagent, the detection main body is a coated 96 orifice plate containing detection thalline, a soda acid indicator and nutrients required for thalline growth, and the detection reagent is an organic solution containing beta-lactam antibiotics. The detection thalline is a bacillus stearothermophilus suspension; the soda acid indicator is bromcresol purple, and the detection reagent is piperacillin dissolved by acetonitrile. The kit is based on a microbe growth inhibition factor releasing method, color change can reflect the microbe growth state, so that the beta-lactamase content case in milk can be indirectly displayed. The kit has the characteristics of simple operation, rapidity, high efficiency and accuracy, and has characteristics of low equipment requirement, no requirement for professional operation, clear result interpretation.

Description

technical field [0001] The invention belongs to the field of food safety detection, and in particular relates to a test kit for β-lactamase added in dairy products and a detection method thereof. Background technique [0002] In recent years, with the rapid development of my country's animal husbandry industry, milk production has increased significantly, and fresh milk and dairy products have become an important part of people's daily food. However, the widespread use of antibiotics in modern animal husbandry inevitably results in antibiotic residues in milk. β-lactamase antibiotics are a kind of spectrum antibiotics derived from microorganisms, which are widely used in the prevention and treatment of livestock and poultry diseases. In order to increase the production of animal food, some are also used as feed additives, which greatly increases the risk of antibiotic residues in food and the environment, causing subsequent human allergic reactions, microbial resistance, pr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/34G01N33/04
Inventor 赵广宇何圣侠
Owner 北京华益精点生物技术有限公司
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