CN107449911A - A kind of tableware flora quick detection collaurum biochip - Google Patents
A kind of tableware flora quick detection collaurum biochip Download PDFInfo
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- CN107449911A CN107449911A CN201710664018.5A CN201710664018A CN107449911A CN 107449911 A CN107449911 A CN 107449911A CN 201710664018 A CN201710664018 A CN 201710664018A CN 107449911 A CN107449911 A CN 107449911A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
- G01N33/56911—Bacteria
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
- G01N33/56911—Bacteria
- G01N33/56916—Enterobacteria, e.g. shigella, salmonella, klebsiella, serratia
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Abstract
The invention particularly discloses a kind of tableware flora quick detection collaurum biochip, including sample layer A, colloid gold label layer B, detection conversion zone C, water accepting layer D and base E, filler T；The filler is absorbent cotton, and the antibody of anti-object bacteria is coated with the colloid gold label layer, detection line M and nature controlling line N are provided with the detection conversion zone.The present invention using colloid gold test paper of the air bladder as membrane structure, there was only a site combined with antigen-specific in gold labeling antibody, there is high specificity；High sensitivity, positive rate basically reach 100%, detection speed is fast, flux is big, simple to operate, colour stable, the advantages that detection temperature optimum range is wide, test paper long shelf-life and cost is cheap, and half-quantitative detection can be realized.The Rapid detection test strip all can be used at 4~50 DEG C, and 5~10min can observe result later, be suitable for the departments such as food and medicine, environmental protection, inspection and quarantine being used for quickly detecting to the gram- bacteria in sample.
Description
Technical field
The present invention relates to technical field of biological, more particularly to a kind of tableware flora quick detection collaurum biology core
Piece.
Background technology
The cleaning of tableware and sterilization method, directly influence the health of household.Bacterium enteritidis
(Salmonellaenteritidis, SE) is the common pathogen for causing food poisoning, has important meaning on public hygienics
Justice, poultry, egg and meat products are SE primary vehicles, drastically influence development and the human health of aquaculture, therefore
To easily strict detection should be carried out before coming into the market by the SE food polluted.
New equipment and detection card emerges in an endless stream, especially in immunology, biochemistry and molecular biology etc. more
It is that have developed various new quick detection detection cards, is improving the detection speed of salmonella, simplifying the numerous of sample treatment
The operation requirement of testing staff is reduced while miscellaneous step, its utilization to detection card is produced dependence.But detection is used
Detection card is widely used in market, and but it is assessed without the reference standard of unified management system and specification, caused
These detections, which are stuck in quality, to be in the presence of uneven, shows sensitivity, specificity and stability difference so that
Testing result cannot ensure occur enteritis in animal food unavoidably in authenticity between each laboratory monitoring/each testing staff
The false retrieval of salmonella or missing inspection, not only it is likely to result in China's inlet and outlet and heavy economic losses occurs in international trade, also
It is possible to threaten to the health of the mankind.And it is current, China is basic in the management that animal food safety is detected on detection card
In blank, for the detection between battle support laboratory while Bacterium enteritidis detection efficiency is improved and between testing staff
Ability, it is ensured that the validity of testing result, it is necessary to use of the specification to detection card product, fill up managerial blank, therefore, grind
Hair can be used for the standard substance of detection card and routine testing quality control imperative.
The content of the invention
For the above-mentioned deficiency of prior art, the present invention provides a kind of tableware flora quick detection collaurum biochip,
To solve above-mentioned technical problem.
The present invention is achieved by the following technical solutions：
A kind of tableware flora quick detection collaurum biochip, including sample layer A, colloid gold label layer B, detection reaction
Layer C, water accepting layer D and base E, filler T；The filler is absorbent cotton, and anti-target is coated with the colloid gold label layer
The antibody of bacterium, described detect are provided with detection line M and nature controlling line N in conversion zone；Object bacteria antigen is coated with the detection line,
The nature controlling line is coated with goat anti-rabbit igg, and the detection gold labeling antibody can be with detection envelope antigen association reaction and showing
Color, and there was only a site combined with antigen-specific in detection gold labeling antibody；The sample layer, colloid gold label layer and inspection
Survey conversion zone to be incorporated in successively on backer board the same face according to order from the inside to the outside, from top to bottom, colloid gold label layer and inspection
The end for surveying conversion zone is overlapping；The water accepting layer is incorporated in the other end of detection conversion zone.
Brief description of the drawings
Structural representation is shown in the side amplification that Fig. 1 is the present invention.
Fig. 2 is the structural representation of the present invention.
Wherein：Sample layer A, colloid gold label layer B, detection conversion zone C, water accepting layer D and base E, filler T, detection line
M, nature controlling line N.
Embodiment
The invention will be further described with the following Examples.
A kind of tableware flora quick detection collaurum biochip, as described in Figure 1, including sample layer A, colloid gold label layer
B, conversion zone C, water accepting layer D and base E, filler T are detected；The filler is absorbent cotton, is wrapped on the colloid gold label layer
The antibody for being had anti-object bacteria, described detect are provided with detection line M and nature controlling line N in conversion zone；It is coated with the detection line
Object bacteria antigen, the nature controlling line are coated with goat anti-rabbit igg, and the detection gold labeling antibody can be with detection envelope antigen knot
Close and react and develop the color, and there was only a site combined with antigen-specific in detection gold labeling antibody；As shown in Fig. 2 the sample
Product layer, colloid gold label layer and detection conversion zone are incorporated on backer board the same face successively according to order from top to bottom, colloid
Golden mark layer is overlapping with the end of detection conversion zone；The water accepting layer is incorporated in the other end of detection conversion zone.
Preferably, the object bacteria includes the one or more in gram-positive bacteria and Gram-negative bacteria.
Preferably, anti-target bacteria antibody is coated with the colloid gold label layer, is using target bacterial outer membrane protein as anti-
Prepared after the immune animal of original.
The strict ratio for controlling colloid gold labeling antibody, object bacteria antigen and goat anti-rabbit igg of the invention., can during concrete application
According to different detection line or parameter line whether develop the color and the depth of color, judge sample liquid in whether contain object bacteria, with
And content is more with less, realizing half-quantitative detection.
Preferably, the object bacteria is Bacterium enteritidis.
Preferably, the detection line is 1~5, and nature controlling line is 1~5.
It is highly preferred that the detection line is 1~3, nature controlling line is 1~3.
Preferably, the sample layer is made up of the good material of water imbibition；The backer board is made up of plastics.
Preferably, the colloid gold label layer and the material of detection conversion zone are biomembrane prepared by air bladder.
Preferably, the colloid gold label layer is biomembrane prepared by black carp, grass carp or silver carp, and the detection conversion zone is
Biomembrane prepared by grass carp, silver carp or bighead.
The present invention is applied in the detection technique of biotechnology using air bladder as biofilm structure first, and air bladder possesses nitric acid
The basic structure of cellulose membrane and some standard biologic films, meets the basic demand of biomembrane, and the source of air bladder is easy, cost
It is cheap, apply it in biological products and a kind of high-quality biomembrane, greatly reduce film part in biological products into
This, increases economic efficiency.
What colloid gold label layer and detection conversion zone in the colloid gold test paper of the present invention were prepared using air bladder for raw material
Biomembrane, air bladder film have good gas permeability and water penetration, cheap and easy to get, high mechanical strength and good toughness, can be enzyme
Immobilization provides excellent bio-matrix.
When two detection lines M1 and M2 show red, then sample is negative (i.e. without object bacteria)；When detection line M1 shows
Color, when detection line M2 does not develop the color, then contain object bacteria in sample, but its concentration is not high；When two detection lines do not develop the color, then
Object bacteria is dense in sample, and half-quantitative detection is carried out with this.
When detection line M colors are deeper than reference line N3, then sample is negative (without object bacteria or denier)；Work as detection line
When M colors are deeper than N3 and are shallower than reference line N2, then sample contains object bacteria, and concentration ratio is relatively low；It is shallower than when detection line M colors are deeper than N2
During N1, then contain object bacteria in sample, concentration is higher；When detection line M does not develop the color, then contain object bacteria, concentration in sample
It is very high, half-quantitative detection is carried out with this.
As a result criterion：
The effect of nature controlling line is：Whether no matter object bacteria is contained in sample solution, and the coated gold mark of colloid gold label layer is anti-
Body moves up can be combined when reaching nature controlling line with the IgG on nature controlling line, and aggregation forms macroscopic red granules, nature controlling line
All the time develop the color, if nature controlling line does not develop the color, represent this test invalidation.
Preferably embodiment, the method that air bladder is applied to biomembrane is will be described in detail below, is comprised the following steps：
S1. the air bladder moisturizing that will be collected into, it is specially that periodically 37 DEG C of deionized waters of spray, subsequent operation process keep air bladder
Keep the state of moistening；
S2. lightly hemispherical air bladder is cut along warp, shakeout, i.e., planarized air bladder；
S3. the air bladder of above-mentioned steps is placed in beaker, adds appropriate amount of deionized water, beaker is placed on oscillator, 46
DEG C, 100~120rpm, 1~2h is vibrated, after vibration terminates, egg shell membrane is placed in the slow water of flowing and rinsed, removes film surface
Impurity；
S4. the air bladder of above-mentioned steps is placed in plane, air bladder is layered outer membrane and inner membrance using tweezers；
S5. outer membrane is placed in 50mMTris-HCl (pH7.5), 10mM CaCl2In solution, it is placed on magnetic stirring apparatus, incubates
40~55 DEG C, 30min are educated, above-mentioned solution adds 20mg/ml Proteinase K 100ul per 100ml, continues 10~20min of incubation；
S6. inner membrance is placed in 50mMTris-HCl (pH7.0), 10mM CaCl2In solution, it is placed on magnetic stirring apparatus, incubates
40~55 DEG C, 10min are educated, above-mentioned solution adds 20mg/ml Proteinase K 10ul per 100ml, continues 5~10min of incubation；
S7. after step S5 and S6 reactions terminate, quickly remove, be placed in 3~5min of flushing in the running water of flowing, make afterwards
With 75% alcohol washes 3~5 times, each 5min；
S8. the film of above-mentioned steps is cleaned 3~5 times, each 3min using deionized water, and 4 DEG C after terminating, moisturizing preserves standby
With.
The present invention using colloid gold test paper of the air bladder as membrane structure, only one and antigen-specific knot in gold labeling antibody
The site of conjunction, there is high specificity；High sensitivity, positive rate basically reach 100%, and detection speed is fast, flux is big, simple to operate
Convenience, colour stable, the advantages that detection temperature optimum range is wide, test paper long shelf-life and cost is cheap, and semidefinite can be realized
Amount detection.The Rapid detection test strip all can be used at 4~50 DEG C, and 5~10min can observe result later, be suitable for food
The departments such as medicine, environmental protection, inspection and quarantine are used for quickly detecting to the gram- bacteria in sample.
Finally it should be noted that the above embodiments are merely illustrative of the technical solutions of the present invention, rather than the present invention is protected
The limitation of scope is protected, although being explained with reference to preferred embodiment to the present invention, one of ordinary skill in the art should
Work as understanding, technical scheme can be modified or equivalent substitution, without departing from the reality of technical solution of the present invention
Matter and scope.
Claims (8)
1. a kind of tableware flora quick detection collaurum biochip, it is characterised in that including sample layer A, colloid gold label layer
B, conversion zone C, water accepting layer D and base E, filler T are detected；The filler is absorbent cotton, is wrapped on the colloid gold label layer
The antibody for being had anti-object bacteria, described detect are provided with detection line M and nature controlling line N in conversion zone；It is coated with the detection line
Object bacteria antigen, the nature controlling line are coated with goat anti-rabbit igg, and the detection gold labeling antibody can be with detection envelope antigen knot
Close and react and develop the color, and there was only a site combined with antigen-specific in detection gold labeling antibody；The sample layer, collaurum
Mark layer and detection conversion zone are incorporated on backer board the same face successively according to order from the inside to the outside, from top to bottom, collaurum
Mark layer is overlapping with the end of detection conversion zone；The water accepting layer is incorporated in the other end of detection conversion zone.
2. tableware flora quick detection collaurum biochip according to claim 1, it is characterised in that the object bacteria
Including the one or more in gram-positive bacteria and Gram-negative bacteria.
3. tableware flora quick detection collaurum biochip according to claim 2, it is characterised in that the collaurum
Anti- target bacteria antibody is coated with mark layer, using target bacterial outer membrane protein as antigen-immunized animal after prepare.
4. tableware flora quick detection collaurum biochip according to claim 3, it is characterised in that the object bacteria
For Bacterium enteritidis.
5. tableware flora quick detection collaurum biochip according to claim 4, it is characterised in that the inspection
Survey line is 1~5, and nature controlling line is 1~5.
6. tableware flora quick detection collaurum biochip according to claim 5, it is characterised in that the sample layer
It is made up of the good material of water imbibition；The backer board is made up of plastics.
7. tableware flora quick detection collaurum biochip according to claim 6, it is characterised in that the collaurum
Mark layer and the material of detection conversion zone are biomembrane prepared by air bladder.
8. tableware flora quick detection collaurum biochip according to claim 7, it is characterised in that the collaurum
Mark layer is biomembrane prepared by black carp, grass carp or silver carp, and the detection conversion zone is biology prepared by grass carp, silver carp or bighead
Film.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710664018.5A CN107449911A (en) | 2017-08-06 | 2017-08-06 | A kind of tableware flora quick detection collaurum biochip |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201710664018.5A CN107449911A (en) | 2017-08-06 | 2017-08-06 | A kind of tableware flora quick detection collaurum biochip |
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| Publication Number | Publication Date |
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| CN107449911A true CN107449911A (en) | 2017-12-08 |
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1458527A (en) * | 2002-08-21 | 2003-11-26 | 中国人民解放军军事医学科学院卫生学环境医学研究所 | Immune chromatographic paper strip and method for quick detecting pathogen and toxin in food |
| CN101031798A (en) * | 2004-07-29 | 2007-09-05 | 瑞莱诊断体系有限公司 | Quantitative lateral flow system and assay |
| CN106093409A (en) * | 2016-06-07 | 2016-11-09 | 江南大学 | The preparation method of the colloidal gold strip that detection Salmonella in Food based on Salmonella core polysaccharide monoclonal antibody belongs to |
| CN106771183A (en) * | 2016-11-29 | 2017-05-31 | 百奥森（江苏）食品安全科技有限公司 | The detection card of Bacterium enteritidis in a kind of detection tableware |
-
2017
- 2017-08-06 CN CN201710664018.5A patent/CN107449911A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1458527A (en) * | 2002-08-21 | 2003-11-26 | 中国人民解放军军事医学科学院卫生学环境医学研究所 | Immune chromatographic paper strip and method for quick detecting pathogen and toxin in food |
| CN101031798A (en) * | 2004-07-29 | 2007-09-05 | 瑞莱诊断体系有限公司 | Quantitative lateral flow system and assay |
| CN106093409A (en) * | 2016-06-07 | 2016-11-09 | 江南大学 | The preparation method of the colloidal gold strip that detection Salmonella in Food based on Salmonella core polysaccharide monoclonal antibody belongs to |
| CN106771183A (en) * | 2016-11-29 | 2017-05-31 | 百奥森（江苏）食品安全科技有限公司 | The detection card of Bacterium enteritidis in a kind of detection tableware |
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Application publication date: 20171208 |