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Method for detecting lactic acid bacteria in probiotic bacteria

A detection method and technology for lactic acid bacteria, which are applied in measurement devices, immunoassays, biological material analysis, etc., can solve the problems of decreased biological activity of lactic acid bacteria, low detection rate of probiotics, long incubation time, etc., and reduce the interference of detection accuracy. , The effect of shortening the detection time and improving the detection rate

Active Publication Date: 2019-03-29
HENAN BUSINESS SCI RES INST
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Problems solved by technology

The traditional national standard detection method takes at least 3 days to produce results, and the cultivation time is long, time-consuming and laborious; in addition, in the actual detection process, the detection environment is not conducive to the growth of bacteria and the long detection time and other factors make the biological activity of lactic acid bacteria decline or even In the case of death, the detection rate of probiotics in the final product is lower than that in the actual sample

Method used

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  • Method for detecting lactic acid bacteria in probiotic bacteria
  • Method for detecting lactic acid bacteria in probiotic bacteria
  • Method for detecting lactic acid bacteria in probiotic bacteria

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preparation example Construction

[0028] Preparation of Lactic Acid Bacteria Immunomagnetic Beads

[0029] a) Dilute the lactic acid bacteria antibody to 1mg / mL with a pH value of 7.4 and a PBST buffer solution with a concentration of 0.01mol / L. The lactic acid bacteria antibody is Bifidobacterium antibody, Streptococcus thermophilus antibody and Lactobacillus antibody in a weight ratio of 4 : 3:3 mixed mixture, prepare 10mg / mL biotin N-hydroxysuccinimide ester solution with anhydrous DMSO, add diluted lactic acid bacteria antibody to the biotin N-hydroxysuccinimide ester solution Mix, the volume ratio of biotin N-hydroxysuccinimide ester solution to lactic acid bacteria antibody is 4:1, and incubate at room temperature to obtain biotin-labeled lactic acid bacteria antibody;

[0030] b) Add 1 mg of activated and dissolved streptavidin-modified magnetic beads to the biotin-labeled lactic acid bacteria antibody for mixing to obtain a mixture of antibody and magnetic beads, and place at room temperature;

[0031...

Embodiment 1

[0032] The selection of embodiment 1 BSAV concentration

[0033] Take 6 parts of 225mL PBS buffer containing different concentrations of BASV (PBS concentration is 0.02mol / L) in homogeneous bags, and add 25g of bifidobacteria, lactobacilli and Streptococcus thermophilus to each homogeneous bag The positive samples were homogenized for 30 seconds. After homogeneous mixing, the sample homogeneous solution was obtained. Since the concentration of the target bacteria in the sample homogeneous solution was high at this time, it could not be counted directly according to the detection method specified in GB4789.35, so the sample needs to be Dilute the homogeneous solution, and dilute the sample homogeneous solution to 10 times according to the 10-fold gradient dilution. -5 , to obtain diluted sample homogenate; the concentrations of BSAV added were 0% (no BSAV added), 0.5%, 1%, 2%, 3% and 5%.

[0034] In addition, take 225mL 0.85% normal saline as the contrast solution, add 25g of ...

Embodiment 2

[0038] Embodiment 2 The selection of lactose solution concentration

[0039] Take 25g of 7 positive samples containing Bifidobacterium, Lactobacillus and Streptococcus thermophilus, add 225mL of PBS buffer solution containing 1% BASV into the homogenization bag, homogenize for 30s, after homogenization, mix according to 10 times Serial dilution, diluted to 10 -5 , to obtain the diluted sample homogeneous solution, add 20mL of lactose solution to the diluted sample homogeneous solution, the concentration of lactose solution is 0% (national standard), 1%, 2%, 5%, 8%, 10% and 20% respectively. %.

[0040]In accordance with the detection method specified in GB4789.35, the above sample liquid was subjected to operations such as enrichment, scribing and separation, and plate counting to obtain the test results. The results are shown in Table 2.

[0041]

[0042] From the results in Table 2, it can be seen that the number of colonies that can be detected is the highest after add...

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Abstract

The invention discloses a method for detecting lactic acid bacteria in probiotic bacteria, and belongs to the technical field of detection for microbial products. The method comprises the following steps: (1) adding a sample to be tested to a PBS buffer solution supplemented with BSAV, homogenizing, and preparing a homogenate of the sample, wherein the ratio of the sample to be tested and the PBSbuffer solution is 1g: 9mL, adding a lactose solution to the homogenate of the sample, and removing a supernatant by a water bath and centrifugation to obtain a bacterial suspension; (2) adding lacticacid bacteria immunomagnetic beads to the bacterial suspension obtained in the step (1), after mixing and shaking, placing the bacterial suspension on a magnetic stand and layering, discarding the upper layer, eluting the lower layer sample, adding the PBS buffer solution to the eluted lactic acid bacteria immunomagnetic beads, and shaking and resuspending magnetic beads to obtain a sample solution to be tested; (3) culturing, observing, and testing the sample solution to be tested. The invention shortens the detection time of the lactic acid bacteria in the sample, improves the detection rate of the lactic acid bacteria and the accuracy of the detection result, and has high detection sensitivity.

Description

technical field [0001] The invention belongs to the technical field of detection of microbial products, and in particular relates to a detection method of lactic acid bacteria in probiotics. Background technique [0002] As a type of active microorganisms that can have beneficial effects on the host's physiological functions, probiotics have the functions of regulating intestinal flora, lowering cholesterol, enhancing immunity, alleviating oral diseases and emotional abnormalities, etc., and are widely used in the fields of food, medicine and Livestock field. In recent years, the research and development, production and sales of lactic acid bacteria and their products in probiotics show that the important functions of lactic acid bacteria in human health have attracted more and more attention from researchers and consumers. Lactic acid bacteria mainly include bifidobacteria , Streptococcus thermophilus and Lactobacillus are more sensitive to ambient temperature. During the ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569
CPCG01N33/56911G01N2333/335G01N2333/315G01N2333/195G01N2469/10
Inventor 周莉王法云刘红伟关炳峰张立攀李向力平洋张亚勋李远远任钊谭静罗蓓蓓王珂王瑞陈俭春
Owner HENAN BUSINESS SCI RES INST
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