CN117761020A - Method and equipment for rapidly detecting bacteria in water body - Google Patents
Method and equipment for rapidly detecting bacteria in water body Download PDFInfo
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- CN117761020A CN117761020A CN202311698052.6A CN202311698052A CN117761020A CN 117761020 A CN117761020 A CN 117761020A CN 202311698052 A CN202311698052 A CN 202311698052A CN 117761020 A CN117761020 A CN 117761020A
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- sample
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- bacteria
- water
- fluorescence intensity
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- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 title claims abstract description 52
- 241000894006 Bacteria Species 0.000 title claims abstract description 39
- 238000000034 method Methods 0.000 title claims abstract description 17
- 238000001514 detection method Methods 0.000 claims abstract description 59
- 238000001914 filtration Methods 0.000 claims abstract description 37
- 230000001580 bacterial effect Effects 0.000 claims abstract description 34
- 239000012528 membrane Substances 0.000 claims abstract description 23
- 238000005070 sampling Methods 0.000 claims abstract description 14
- 239000006166 lysate Substances 0.000 claims abstract description 13
- 239000000376 reactant Substances 0.000 claims abstract description 11
- 238000006243 chemical reaction Methods 0.000 claims abstract description 8
- 239000000706 filtrate Substances 0.000 claims abstract description 8
- 230000002452 interceptive effect Effects 0.000 claims abstract description 8
- 239000000203 mixture Substances 0.000 claims abstract description 7
- 239000000463 material Substances 0.000 claims description 11
- 239000011148 porous material Substances 0.000 claims description 11
- 239000002184 metal Substances 0.000 claims description 9
- 229910052751 metal Inorganic materials 0.000 claims description 9
- 239000000725 suspension Substances 0.000 claims description 7
- 108060001084 Luciferase Proteins 0.000 claims description 5
- 239000005089 Luciferase Substances 0.000 claims description 5
- 230000002572 peristaltic effect Effects 0.000 claims description 5
- IGXWBGJHJZYPQS-SSDOTTSWSA-N D-Luciferin Chemical compound OC(=O)[C@H]1CSC(C=2SC3=CC=C(O)C=C3N=2)=N1 IGXWBGJHJZYPQS-SSDOTTSWSA-N 0.000 claims description 3
- CYCGRDQQIOGCKX-UHFFFAOYSA-N Dehydro-luciferin Natural products OC(=O)C1=CSC(C=2SC3=CC(O)=CC=C3N=2)=N1 CYCGRDQQIOGCKX-UHFFFAOYSA-N 0.000 claims description 3
- BJGNCJDXODQBOB-UHFFFAOYSA-N Fivefly Luciferin Natural products OC(=O)C1CSC(C=2SC3=CC(O)=CC=C3N=2)=N1 BJGNCJDXODQBOB-UHFFFAOYSA-N 0.000 claims description 3
- DDWFXDSYGUXRAY-UHFFFAOYSA-N Luciferin Natural products CCc1c(C)c(CC2NC(=O)C(=C2C=C)C)[nH]c1Cc3[nH]c4C(=C5/NC(CC(=O)O)C(C)C5CC(=O)O)CC(=O)c4c3C DDWFXDSYGUXRAY-UHFFFAOYSA-N 0.000 claims description 3
- 239000007800 oxidant agent Substances 0.000 claims description 3
- 239000003651 drinking water Substances 0.000 description 7
- 235000020188 drinking water Nutrition 0.000 description 7
- 238000012360 testing method Methods 0.000 description 5
- GNBHRKFJIUUOQI-UHFFFAOYSA-N fluorescein Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 GNBHRKFJIUUOQI-UHFFFAOYSA-N 0.000 description 4
- 238000005374 membrane filtration Methods 0.000 description 4
- 238000012136 culture method Methods 0.000 description 3
- 238000010586 diagram Methods 0.000 description 3
- 238000001917 fluorescence detection Methods 0.000 description 3
- 241000700605 Viruses Species 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 238000012258 culturing Methods 0.000 description 2
- LIYGYAHYXQDGEP-UHFFFAOYSA-N firefly oxyluciferin Natural products Oc1csc(n1)-c1nc2ccc(O)cc2s1 LIYGYAHYXQDGEP-UHFFFAOYSA-N 0.000 description 2
- 239000001963 growth medium Substances 0.000 description 2
- 238000001471 micro-filtration Methods 0.000 description 2
- JJVOROULKOMTKG-UHFFFAOYSA-N oxidized Photinus luciferin Chemical compound S1C2=CC(O)=CC=C2N=C1C1=NC(=O)CS1 JJVOROULKOMTKG-UHFFFAOYSA-N 0.000 description 2
- 150000003384 small molecules Chemical class 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 description 1
- 229910052802 copper Inorganic materials 0.000 description 1
- 239000010949 copper Substances 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 230000001590 oxidative effect Effects 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 238000011282 treatment Methods 0.000 description 1
Abstract
The invention discloses a method and equipment for rapidly detecting bacteria in a water body, wherein the method comprises the following steps: sampling to be detected to obtain a sample to be detected; taking a certain volume of sample to be detected, and carrying out microporous filtration to obtain a filtrate; collecting the filtered matter, mixing the filtered matter with the lysate and fluorescent reactant, and detecting the relative fluorescence intensity of the mixture; measuring and calculating the bacterial number in a sample to be detected according to the relative fluorescence intensity; the rapid detection equipment for the bacteria in the water body comprises a filtering system and a detection system; the filtration system comprises a sample container, a filter and a power pump; the detection system comprises a collection device, a photoelectric conversion unit, a control module and an interactive display module. After the water sample is filtered, residues on the filtering membrane are collected by using an integrated swab, and then the fluorescence intensity of the swab part is detected by using a photoelectric detection unit, and according to the fluorescence intensity and the volume of the filtered water sample, the bacterial pollution condition in the detected water body, namely whether the bacterial content exceeds the standard or not and the value interval of the bacterial content can be judged.
Description
Technical Field
The invention relates to the technical field of detection methods and equipment, in particular to a method and equipment for rapidly detecting bacteria in a water body.
Background
The existing bacteria detection method mainly comprises a membrane filtration culture method and an ATP rapid detection method, wherein bacteria in a sample are filtered and trapped by a filter membrane and then placed into a culture medium for culturing for 24 to 48 hours, then bacterial colonies are observed, counted and other subsequent detection are carried out, the detection result is more accurate, the types and the quantity of the bacteria can be detected, but the time consumption is longer, and the method is not suitable for occasions needing on-site rapid detection; the latter mainly uses the ATP content in living bacterial cells to be in direct proportion to the bacterial quantity, under the condition of ATP existence, luciferase catalyzes ATP to react with luciferin to generate oxyluciferin and emit visible light, and the light intensity can reflect the ATP content, thereby indirectly reflecting the bacterial content in a sample; the detection process can only perform qualitative detection due to the limitation of the sampling mode, and for a water body (the nature of which is bacterial suspension) possibly polluted, such as drinking water in a drinking water system, the sampling mode of simply adopting a swab to scrape cannot detect a corresponding result with great probability, and detection omission can be caused when serious.
Disclosure of Invention
In view of the above-mentioned shortcomings of the current culture method for detecting bacteria and ATP fluorescence detection technology, the invention provides a rapid detection method and device for water bacteria, which can be achieved.
In order to achieve the above purpose, the embodiment of the present invention adopts the following technical scheme:
a method for rapidly detecting bacteria in a water body is characterized by comprising the following steps:
sampling to be detected to obtain a sample to be detected;
taking a certain volume of sample to be detected, and carrying out microporous filtration to obtain a filtrate;
collecting the filtered matter, mixing the filtered matter with the lysate and fluorescent reactant, and detecting the relative fluorescence intensity of the mixture;
and measuring and calculating the bacterial quantity in the sample to be detected according to the relative fluorescence intensity.
According to one aspect of the invention, the pore size used in the microfiltration of a volume of sample to be tested to obtain the filtrate is 0.45 μm or less, and the pore size is set to ensure that most bacteria are trapped and water molecules and other small molecules pass through.
According to one aspect of the invention, the fluorescent reactant comprises fluorescein, luciferase, an oxidizing agent, and a buffer.
According to one aspect of the invention, the measuring the bacterial count in the sample to be detected based on the relative fluorescence intensity comprises:
taking bacterial suspension with known total bacteria number, and carrying out microporous filtration to obtain a standard filter material;
collecting standard filter materials, mixing the standard filter materials with lysate and fluorescent reactant, and detecting the fluorescence intensity of the mixture; the fluorescence intensity was used as a reference for comparison.
A rapid detection device for bacteria in a water body comprises a filtering system and a detection system; the filtration system includes a sample container, a filter, and a powered pump; the detection system comprises a collection device, a photoelectric conversion unit, a control module and an interactive display module; one side of the filter is connected with the sample container, and the other side of the filter is connected with the power pump through a pipeline; the photoelectric conversion unit and the interactive display module are respectively and electrically connected with the control module.
According to one aspect of the invention, the filter is a filtering membrane and a metal filter core in sequence from top to bottom; wherein filtering membrane tiling is placed in metal filter core top, and metal filter core is porous structure, mainly plays and supports filtering membrane, lets when filtering, the balanced effect of pressure of filtering membrane everywhere.
According to one aspect of the invention, the pore size of the filtering membrane is 0.45 μm or less, and the pore size can intercept most of bacterial cells known at present and allow water molecules and other small molecular substances in water to pass through, so that the accuracy of detection results is ensured.
According to one aspect of the invention, the collection device is a swab; the swab head of the swab contains a lysate; the swab adopts an integrated swab, namely, reagent components including fluorescein, lysate and the like are designed into a whole with the swab, so that the purposes of convenient carrying and convenient detection and use are achieved.
According to one aspect of the invention, the power pump is a peristaltic pump; peristaltic pump simple structure can be miniaturized, can improve equipment portability.
The implementation of the invention has the advantages that:
through combining the membrane filtration technology with the ATP detection technology, use the filtration membrane of micropore to accurately filter and hold back bacteria, then adopt ATP fluorescence detection means to detect its fluorescence intensity, compare more quick in culture detection method, only need ten minutes from the sampling to testing result come out, compare in traditional ATP fluorescence rapid detection, its testing result is more accurate, and because can filter and detect a large amount of water samples, be more suitable for carrying out on-the-spot rapid determination analysis to drinking water, a large amount of water bodies of domestic water, avoid sampling point, the leak detection that the sampling volume is too little to cause.
Drawings
In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings that are needed in the embodiments will be briefly described below, and it is obvious that the drawings in the following description are only some embodiments of the present invention, and other drawings may be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 is a block flow diagram of a method for rapidly detecting bacteria in a body of water according to an embodiment of the present invention;
FIG. 2 is a block diagram of a detection system according to an embodiment of the present invention;
FIG. 3 is a schematic structural diagram of a rapid detection device for bacteria in a water body according to an embodiment of the present invention;
fig. 4 is a perspective structural view of a rapid water bacteria detection device according to an embodiment of the present invention.
Legend description: 1. a sample container; 2. a filter; 21. a filtering membrane; 22. a metal filter element; 3. a power pump; 4. a water outlet; 5. a swab; 6. a photoelectric conversion unit; 7. a control module; 8. an electronic screen; 9. and inputting keys.
Detailed Description
The following description of the embodiments of the present invention will be made clearly and completely with reference to the accompanying drawings, in which it is apparent that the embodiments described are only some embodiments of the present invention, but not all embodiments. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
The existing bacterial detection method aiming at the water body mainly comprises a membrane filtration culture method and an ATP rapid detection method, wherein bacteria in a sample are filtered and trapped by a filter membrane and then are put into a culture medium for culturing for 24 to 48 hours, then bacterial colonies are observed, counted and other subsequent detection are carried out, the detection result is more accurate, the types and the quantity of the bacteria can be detected, but the time consumption is longer, and the method is not suitable for occasions needing on-site rapid detection;
the existing ATP rapid detection method mainly utilizes the fact that the ATP content in living bacterial cells is in direct proportion to the bacterial quantity, under the condition that ATP exists, luciferase catalyzes ATP to react with luciferin to generate oxyluciferin and emit visible light, the ATP content can be reflected by the light intensity, and therefore the bacterial content in a sample is indirectly reflected; the common ATP fluorescence detector adopts an integrated swab to sample the surface of an object to be detected, and detects the fluorescence intensity by using a matched fluorescence detector, thereby rapidly judging whether bacteria residues exist on the surface of the object to be detected, and the detection process can only carry out qualitative detection due to the limitation of the sampling mode, and the sampling mode has high probability that corresponding results can not be detected for water bodies possibly polluted, such as drinking water in a drinking water system.
As shown in fig. 1, 2, 3 and 4, a method for rapidly detecting bacteria in a water body is characterized by comprising the following steps:
s1, sampling to be detected to obtain a sample to be detected;
s2, taking a certain volume of sample to be detected, and carrying out microporous filtration to obtain a filtrate;
s3, collecting the filter material, mixing the filter material with the lysate and the fluorescent reactant, and detecting the relative fluorescence intensity of the mixture;
and S4, measuring and calculating the number of bacteria in the sample to be detected according to the relative fluorescence intensity.
And step S2, carrying out microporous filtration on a certain volume of sample to be detected to obtain a filtrate, wherein the filter pore diameter adopted in the filtrate is 0.45 mu m or less, and the pore diameter can ensure that most bacteria are trapped and water molecules and other small molecules pass through.
The lysate in step S3 is used to destroy bacterial cells so that ATP therein can react with fluorescent reactants; wherein the fluorescent reactant comprises fluorescein, luciferase, an oxidant and a buffer.
In practical applications, the measuring the bacterial count in the sample to be detected according to the relative fluorescence intensity in step S4 includes:
taking bacterial suspension with known total bacteria number, and carrying out microporous filtration to obtain a standard filter material;
collecting standard filter materials, mixing the standard filter materials with lysate and fluorescent reactant, and detecting the fluorescence intensity of the mixture; taking the fluorescence intensity as a reference to participate in comparison; in practical application, bacterial suspensions with determined bacterial concentration can be prepared in advance, a plurality of bacterial suspensions with different concentration levels are obtained after gradient dilution, the corresponding fluorescence intensities under different bacterial concentrations are obtained through the steps respectively, and the range of the bacterial concentration in a water sample to be detected can be measured more accurately as a reference in actual detection.
A rapid detection device for bacteria in a water body comprises a filtering system and a detection system; the filtration system comprises a sample container 1, a filter 2 and a power pump 3; the detection system comprises a collection device, a photoelectric conversion unit 6, a control module 7 and an interactive display module; one side of the filter 2 is connected with the sample container 1, and the other side is connected with the power pump 3 through a pipeline; the photoelectric conversion unit 6 and the interactive display module are respectively and electrically connected with the control module 7; in practical application, the interactive display module mainly comprises an electronic screen 8 for displaying detection results and an input key 9 for interaction; the control module can be preset with fluorescence intensity data obtained by detecting bacterial suspensions with different concentration levels to serve as a reference for comparison, and the data can be calibrated through a later test.
The filter 2 is provided with a filtering membrane 21 and a metal filter element 22 from top to bottom in sequence; because the filtering membrane 21 is generally thinner and has lower strength, and is difficult to bear larger water pressure under the condition of no support, the filtering membrane 21 is flatly laid above the metal filter element 22, and the metal filter element 22 is of a porous structure and mainly plays a role in supporting the filtering membrane 21, so that the pressure of the filtering membrane 21 is balanced when in filtering; the practical application adopts a porous copper filter element.
The pore diameter of the filtering membrane 21 is 0.45 μm or less, and most of bacterial cells known at present can be trapped by the pore diameter, so that water molecules and other small molecular substances in the water can pass through the membrane, and the accuracy of a detection result is ensured.
The collecting device is a swab 5; the swab 5 head of the swab 5 contains a lysate; the swab 5 adopts an integrated swab 5, namely, reagent components comprising fluorescein, lysate and the like are designed as a whole with the swab 5, so that the purposes of convenient carrying, convenient detection and use are achieved.
The power pump 3 is a peristaltic pump; peristaltic pump simple structure can be miniaturized, can improve equipment portability.
According to the technical scheme, the method is mainly used for detecting whether bacteria are polluted in domestic water such as drinking water or not, so that the sampled water sample is subjected to various treatments in advance and can be directly detected, when the method is used, a certain amount of water sample to be detected is injected into a sample container 1, a power valve is opened, suction generated by the power valve is used for sucking the water sample to pass through a filter 2 so that bacterial cells in the water sample are trapped by a filter membrane 21, an integrated swab 5 is used for collecting residues on the filter membrane 21, then a photoelectric detection unit is used for detecting the fluorescence intensity of a swab 5 part, and the bacterial pollution condition in the detected water body, namely whether the bacterial content exceeds a standard or not and the value interval of the bacterial content can be judged according to the fluorescence intensity and the volume of the filtered water sample; further, a membrane with smaller pore diameter parameter is replaced, so that specific viruses can be filtered and trapped, and whether the water sample contains residues of the specific viruses can be detected by adopting a corresponding detection swab; similarly, for any detection target that can be separated from water by microfiltration and has a corresponding rapid detection means, a similar procedure can be used for detection.
The implementation of the invention has the advantages that:
through combining the membrane filtration technology with the ATP detection technology, use the filtration membrane of micropore to accurately filter and hold back bacteria, then adopt ATP fluorescence detection means to detect its fluorescence intensity, compare more quick in culture detection method, only need ten minutes from the sampling to testing result come out, compare in traditional ATP fluorescence rapid detection, its testing result is more accurate, and because can filter and detect a large amount of water samples, be more suitable for carrying out on-the-spot rapid determination analysis to drinking water, a large amount of water bodies of domestic water, avoid sampling point, the leak detection that the sampling volume is too little to cause.
The foregoing is merely illustrative of the present invention, and the present invention is not limited thereto, and any changes or substitutions easily contemplated by those skilled in the art within the technical scope of the present invention should be included in the scope of the present invention. Therefore, the protection scope of the present invention shall be subject to the protection scope of the claims.
Claims (9)
1. A method for rapidly detecting bacteria in a water body is characterized by comprising the following steps:
sampling to be detected to obtain a sample to be detected;
taking a certain volume of sample to be detected, and carrying out microporous filtration to obtain a filtrate;
collecting the filtered matter, mixing the filtered matter with the lysate and fluorescent reactant, and detecting the relative fluorescence intensity of the mixture;
and measuring and calculating the bacterial quantity in the sample to be detected according to the relative fluorescence intensity.
2. The rapid detection method of water bacteria according to claim 1, wherein the filtration pore size adopted in the filtrate obtained by taking a certain volume of the sample to be detected and carrying out microporous filtration is 0.45 μm or less.
3. The rapid detection method of water bacteria of claim 1, wherein the fluorescent reactant comprises luciferin, luciferase, an oxidizing agent and a buffer.
4. The method for rapidly detecting bacteria in a water body according to claim 1, wherein the measuring the number of bacteria in the sample to be detected based on the relative fluorescence intensity comprises:
taking bacterial suspension with known total bacteria number, and carrying out microporous filtration to obtain a standard filter material;
collecting standard filter materials, mixing the standard filter materials with lysate and fluorescent reactant, and detecting the fluorescence intensity of the mixture; the fluorescence intensity was used as a reference for comparison.
5. The rapid detection equipment for the bacteria in the water body is characterized by comprising a filtering system and a detection system; the filtration system includes a sample container, a filter, and a powered pump; the detection system comprises a collection device, a photoelectric conversion unit, a control module and an interactive display module; one side of the filter is connected with the sample container, and the other side of the filter is connected with the power pump through a pipeline; the photoelectric conversion unit and the interactive display module are respectively and electrically connected with the control module.
6. The rapid detection device for water bacteria according to claim 5, wherein the filter comprises a filtering membrane and a metal filter core from top to bottom in sequence; the metal filter element is of a porous structure.
7. The rapid detection apparatus for water bacteria according to claim 6, wherein the pore diameter of the filtering membrane is 0.45 μm or less.
8. The rapid detection apparatus for water bacteria of claim 5, wherein the collection device is a swab; the swab head of the swab contains a lysate.
9. The rapid detection apparatus for water bacteria of claim 5, wherein the power pump is a peristaltic pump.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311698052.6A CN117761020A (en) | 2023-12-12 | 2023-12-12 | Method and equipment for rapidly detecting bacteria in water body |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311698052.6A CN117761020A (en) | 2023-12-12 | 2023-12-12 | Method and equipment for rapidly detecting bacteria in water body |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN117761020A true CN117761020A (en) | 2024-03-26 |
Family
ID=90323102
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202311698052.6A Pending CN117761020A (en) | 2023-12-12 | 2023-12-12 | Method and equipment for rapidly detecting bacteria in water body |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN117761020A (en) |
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2023
- 2023-12-12 CN CN202311698052.6A patent/CN117761020A/en active Pending
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