CN111983212A - Centrifugal microfluidic detection system and method for seminal plasma fructose - Google Patents
Centrifugal microfluidic detection system and method for seminal plasma fructose Download PDFInfo
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- CN111983212A CN111983212A CN202010703621.1A CN202010703621A CN111983212A CN 111983212 A CN111983212 A CN 111983212A CN 202010703621 A CN202010703621 A CN 202010703621A CN 111983212 A CN111983212 A CN 111983212A
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Abstract
The invention provides a centrifugal microfluidic detection system and method for seminal plasma fructose, wherein the method comprises the following steps: s1, mixing the seminal plasma, the diluent and the deproteinized liquid, injecting the mixture into a sample injection cavity, and fully dissolving and reacting the mixture with alkaline solids to form a sample; s2, starting the centrifugal machine to rotate the microfluidic chip, enabling the prepackaged reagent in the central reagent cavity to enter the reagent quantitative cavity, and enabling the sample in the sample injection cavity to enter the separation quantitative cavity; s3, starting the centrifugal machine to rotate the microfluidic chip, enabling the sample in the separation quantitative cavity and the prepackaged reagent in the reagent quantitative cavity to respectively enter the mixing cavity, and dissolving the indole freeze-drying balls in the mixing cavity to form a mixed solution; and S4, starting the centrifugal machine to rotate the microfluidic chip, so that the mixed liquid enters the test cavity and is mixed with the protective agent freeze-drying balls in the test cavity. Compared with the traditional manual operation method, the method provided by the invention can greatly simplify the test flow, reduce the influence of people on the detection result in the test process, and improve the detection efficiency and the reliability of the detection result.
Description
Technical Field
The invention relates to the technical field of seminal plasma fructose detection, in particular to a centrifugal microfluidic detection system and method for seminal plasma fructose.
Background
Fructose in seminal plasma is secreted by the seminal vesicle, influenced by testosterone levels in the blood, and is a major source of sperm energy. Currently, the determination of fructose content is commonly used for monitoring the function of seminal vesicle and testicular interstitial cells and for the diagnosis of infertility. The recommended detection method of the world health organization is an indole method, and the specific process is as follows: 1) adding the seminal plasma into the diluent and the deproteinizing solution for mixing; 2) adding an alkaline solution and mixing; 3) standing at room temperature, and starting a centrifugal machine for centrifugation; 4) adding indole reagent and concentrated hydrochloric acid and mixing; 5) heating in water bath for a certain time, and cooling for a certain time; 6) a sample injector samples a certain amount of mixed solution into the enzyme-labeled micropores; 7) adding a protective agent in a certain proportion; 8) and (5) colorimetric reading of a microplate reader. The whole process needs to use various reagents including protein removing liquid, alkaline solution, diluent, protective agent, concentrated hydrochloric acid and the like, needs operations such as centrifugation by a centrifugal machine, heating by a water bath box, color development of an enzyme-labeling instrument and the like, is complex and tedious, and is completed by manual operation.
Because the whole process of the indole method is complicated, the manual operation is time-consuming and labor-consuming, has high requirements on operators and needs to have related professional skills. Meanwhile, under the influence of people, detection results of different laboratories have wide difference. Aiming at the defects of the manual operation detection process, the invention provides a full-automatic detection method of seminal plasma fructose based on the existing centrifugal microfluidic chip system and the specific process of an indole method.
Disclosure of Invention
In order to solve the problems that the seminal plasma fructose detection manual operation is complicated in process and difficult to ensure consistent results, the invention provides a seminal plasma fructose centrifugal microfluidic detection system and a seminal plasma fructose centrifugal microfluidic detection method, so that the whole test flow is simplified, automatic detection is realized, the influence on the results caused by human factors in the manual operation process is avoided, and the reliability of the detection results is effectively improved.
In order to achieve the purpose, the invention adopts the following specific technical scheme:
the invention provides a centrifugal type seminal plasma fructose microfluidic detection system, which comprises a centrifugal machine and a centrifugal type seminal plasma fructose analysis microfluidic chip arranged on a main shaft of the centrifugal machine, wherein the centrifugal type seminal plasma fructose analysis microfluidic chip comprises a central reagent cavity packaged with a pre-packaged reagent, a sample injection cavity packaged with alkaline solids, a separation quantitative cavity, a reagent quantitative cavity, a mixing cavity packaged with an indole freeze-drying ball and at least one test cavity packaged with a protective agent freeze-drying ball; the central reagent cavity is communicated with the reagent quantifying cavity through a channel, the sample feeding cavity is communicated with the separating quantifying cavity through a channel, the separating quantifying cavity and the reagent quantifying cavity are respectively communicated with the mixing cavity through channels, and the mixing cavity is communicated with the testing cavity through a channel.
Preferably, the sample waste chamber is in communication with the separation quantification chamber via a channel.
Preferably a reagent waste chamber communicating with the reagent dosing chamber via a channel.
Preferably a mixed liquor waste chamber in communication with the test chamber via a channel.
Preferably, the pre-encapsulation reagent is concentrated hydrochloric acid.
Preferably, the alkaline solid is a sodium hydroxide solid.
The invention also provides a centrifugal microfluidic detection method for seminal plasma fructose, which comprises the following steps:
s1, mixing the seminal plasma, the diluent and the deproteinized liquid, injecting the mixture into a sample injection cavity, and fully dissolving and reacting the mixture with alkaline solids to form a sample;
s2, starting a centrifugal machine to rotate the centrifugal type seminal plasma fructose analysis microfluidic chip, enabling a pre-packaged reagent in the central reagent cavity to enter a reagent quantitative cavity, and enabling a sample in the sample injection cavity to enter a separation quantitative cavity;
s3, starting a centrifugal machine to rotate the centrifugal type seminal plasma fructose analysis microfluidic chip, enabling the sample in the separation quantitative cavity and the prepackaged reagent in the reagent quantitative cavity to respectively enter a mixing cavity, and dissolving the indole freeze-drying balls in the mixing cavity to form a mixed solution;
and S4, starting the centrifugal machine to rotate the centrifugal seminal plasma fructose analysis microfluidic chip, enabling the mixed liquid to enter the test cavity and mixing with the protective agent freeze-drying balls in the test cavity.
The invention can obtain the following technical effects:
the automation of the indole fructose detection process can be realized by adopting the modes of reagent prepackaging and centrifugal microfluidic driving, compared with the traditional manual operation method, the method can greatly simplify the test flow, reduce the influence of people on the detection result in the test process, and improve the reliability of the detection result while improving the detection efficiency.
Drawings
FIG. 1 is a schematic diagram of the structure of a centrifugal microfluidic detection system for fructose syrup according to an embodiment of the present invention;
fig. 2 is a schematic flow diagram of a centrifugal microfluidic detection method for seminal fructose according to an embodiment of the invention.
Wherein the reference numerals include: centrifugal seminal plasma fructose analysis micro-fluidic chip 1, central reagent chamber 2, advance kind chamber 3, separation ration chamber 4, reagent ration chamber 5, mixing chamber 6, test chamber 7, prepackage reagent 8, alkaline solid 9, indole freeze-drying ball 10, protectant freeze-drying ball 11, sample waste liquid chamber 12, reagent waste liquid chamber 13, mixed liquid waste liquid chamber 14.
Detailed Description
Hereinafter, embodiments of the present invention will be described with reference to the accompanying drawings. In the following description, the same reference numerals are used for the same blocks. In the case of the same reference numerals, their names and functions are also the same. Therefore, detailed description thereof will not be repeated.
In order to make the objects, technical solutions and advantages of the present invention more apparent, the present invention will be described in further detail below with reference to the accompanying drawings and specific embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not to be construed as limiting the invention.
Specific embodiments of the present invention will be described in detail below with reference to the accompanying drawings.
Fig. 1 shows the structure of a centrifugal microfluidic detection system for fructose syrup according to one embodiment of the present invention.
As shown in fig. 1, the centrifugal microfluidic detection system for fructose syrup provided by the embodiment of the present invention includes: the centrifugal type seminal plasma fructose analysis micro-fluidic chip 1 is arranged on a main shaft of the centrifugal machine, and the centrifugal type seminal plasma fructose analysis micro-fluidic chip 1 is driven by the centrifugal machine to rotate so as to generate centrifugal force.
Centrifugal seminal plasma fructose analysis micro-fluidic chip 1 includes: the device comprises a central reagent cavity 2, a sample injection cavity 3, a separation quantitative cavity 4, a reagent quantitative cavity 5, a mixing cavity 6 and a test cavity 7; wherein, the central reagent cavity 2 is internally sealed with a pre-packaged reagent 8, for example, the pre-packaged reagent 8 is a corrosive strong acid solution such as concentrated hydrochloric acid, etc., so as to avoid over-light color development.
The reagent quantitative cavity 5 is communicated with the central reagent cavity 2 through a channel, and when the centrifugal type seminal plasma fructose analysis micro-fluidic chip 1 rotates, the pre-packaged reagent 8 flows into the reagent quantitative cavity 5 from the central reagent cavity 2 through the generated centrifugal force.
The sample cavity 3 is sealed with alkaline solid 9, such as alkaline solid reagent such as sodium hydroxide solid, and the alkaline solid reagent is injected into the sample cavity 3 after the seminal plasma, the diluent and the deproteinized liquid are mixed, and stands until the alkaline solid 9 is fully dissolved and reacts to form a sample for detection.
The separation quantitative cavity 4 is communicated with the sample injection cavity 3 through a channel, and when the centrifugal type seminal plasma fructose analysis micro-fluidic chip 1 rotates, a sample flows into the separation quantitative cavity 4 from the sample injection cavity 3 through the generated centrifugal force.
The separation quantitative cavity 4 and the reagent quantitative cavity 5 are respectively communicated with the mixing cavity 6 through channels, and an indole freeze-drying ball 10 (freeze-drying indole to form a solid small ball) is sealed in the mixing cavity 6. When the centrifugal type seminal plasma fructose analysis micro-fluidic chip 1 rotates, under the action of centrifugal force, the sample in the separation quantitative cavity 4 and the pre-packaged reagent 8 in the reagent quantitative cavity 5 respectively flow into the mixing cavity 6 to dissolve the indole freeze-dried balls 10 in the mixing cavity 6, so as to form mixed liquid.
The number of the test cavities 7 is at least one, each test cavity 7 is communicated with the mixing cavity 6 through a channel in sequence, and each test cavity 7 is internally and respectively packaged with a protective agent freeze-drying ball 11 (the protective agent is freeze-dried to form a solid small ball). When the centrifugal type seminal plasma fructose analysis micro-fluidic chip 1 rotates, under the action of centrifugal force, the mixed liquid flows into the testing cavity 7 from the mixing cavity 6 and is mixed with the protective agent freeze-drying balls 11 in the testing cavity 7.
Through encapsulating alkaline solid 9 in advancing kind chamber 3, encapsulate indole freeze-drying ball 10 in mixing chamber 6 to and encapsulate protectant freeze-drying ball 11 in test chamber 7, can reduce the ration of detect in-process reagent and add the operation, reduce the influence of test process people to the testing result, improve the credibility of testing result.
In an embodiment of the present invention, the centrifugal microfluidic chip 1 for analyzing fructose contained in seminal plasma further comprises a sample waste chamber 12, wherein the sample waste chamber 12 is communicated with the separation quantification chamber 4 through a channel, and after the separation quantification chamber 4 is filled, the excessive sample overflows and flows into the sample waste chamber 12.
In another embodiment of the invention, the centrifugal type seminal plasma fructose analysis microfluidic chip 1 further comprises a reagent waste liquid cavity 13, the reagent waste liquid cavity 13 is communicated with the reagent quantifying cavity 5 through a channel, and after the reagent quantifying cavity 5 is filled, the redundant prepackaged reagent 8 can overflow and flow into the reagent waste liquid cavity 13.
In another embodiment of the present invention, the centrifugal type seminal plasma fructose analysis microfluidic chip 1 further comprises a mixed liquid waste chamber 14, the mixed liquid waste chamber 14 is communicated with the test chambers 7 through a channel, and after all the test chambers 7 are filled, the redundant mixed liquid overflows the last test chamber 7 and flows into the mixed liquid waste chamber 14.
The above details explain the structure and the working principle of the centrifugal microfluidic detection system for fructose syrup provided by the present invention. Corresponding to the centrifugal microfluidic detection system for the seminal fructose, the invention also provides a method for detecting the seminal fructose by using the centrifugal microfluidic detection system for the seminal fructose.
Fig. 2 shows a flow of a centrifugal microfluidic detection method of seminal fructose according to an embodiment of the present invention.
As shown in fig. 2, the centrifugal microfluidic detection method for fructose in seminal plasma provided by the embodiment of the invention comprises the following steps:
and S1, mixing the seminal plasma, the diluent and the deproteinized liquid, injecting the mixture into the sample injection cavity, and fully dissolving and reacting the mixture with the alkaline solid to form a sample.
The alkaline solid can be alkaline solid reagents such as sodium hydroxide, etc., and is injected into the sample injection cavity for standing after mixing the seminal plasma, the diluent and the deproteinized liquid, so that the sodium hydroxide solid is fully dissolved and reacted. Thereby forming a sample for detection.
And S2, starting the centrifugal type seminal plasma fructose analysis microfluidic chip of the centrifugal machine to rotate, enabling the prepackaged reagent in the central reagent cavity to enter the reagent quantitative cavity, and enabling the sample in the sample injection cavity to enter the separation quantitative cavity.
The centrifuge is started and the centrifugal seminal plasma fructose analysis microfluidic chip is rotated at 5000 rpm for 20 minutes to allow the prepackaged reagent (e.g. concentrated hydrochloric acid) to flow from the central reagent chamber into the reagent dosing chamber and the sample to flow from the sample injection chamber into the separation dosing chamber.
And S3, starting the centrifugal machine to rotate the centrifugal type seminal plasma fructose analysis microfluidic chip, enabling the sample in the separation quantitative cavity and the concentrated hydrochloric acid in the reagent quantitative cavity to respectively enter the mixing cavity, and dissolving the indole freeze-drying balls in the mixing cavity to form a mixed solution.
And starting the centrifuge at an angular acceleration of 2000 revolutions per minute (seconds), and continuing for 30 seconds after the acceleration reaches 2000 revolutions per minute, so that the sample in the separation quantitative cavity and the pre-packaged reagent in the reagent quantitative cavity flow into the mixing cavity to dissolve the indole freeze-dried balls in the mixing cavity to form a mixed solution.
In one embodiment of the invention, the centrifuge is started to accelerate to 3000 rpm at 3000 rpm/(min · s) and then decelerated to a stop for 5 cycles to complete the mixing of the sample, concentrated hydrochloric acid and indole in the mixing chamber.
And S4, starting the centrifugal machine to rotate the centrifugal seminal plasma fructose analysis microfluidic chip, enabling the mixed liquid to enter the test cavity and mixing with the protective agent freeze-drying balls in the test cavity.
The centrifuge was started at an angular acceleration of 6000 revolutions per minute (ms), and after an acceleration to 6000 rpm, which lasted for 30 seconds, the mixed liquid was flowed into the test chamber and mixed with the protectant in the test chamber.
The automatic detection is completed by controlling the driving process of the centrifuge at different rotating speeds through a program, and no additional manual operation is needed in the steps except for the steps S1 and S2. Before step S7, the temperature is 50 ℃ for 20 minutes, the temperature is 0 ℃ for 15 minutes, and the process is completed by a temperature control system on a chip, and the final colorimetric process is completed by a photometric system in a test chamber according to the description of the indole method.
In the description herein, references to the description of the term "one embodiment," "some embodiments," "an example," "a specific example," or "some examples," etc., mean that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the invention. In this specification, the schematic representations of the terms used above are not necessarily intended to refer to the same embodiment or example. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples. Furthermore, various embodiments or examples and features of different embodiments or examples described in this specification can be combined and combined by one skilled in the art without contradiction.
Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention, and that variations, modifications, substitutions and alterations can be made to the above embodiments by those of ordinary skill in the art within the scope of the present invention.
The above embodiments of the present invention should not be construed as limiting the scope of the present invention. Any other corresponding changes and modifications made according to the technical idea of the present invention should be included in the protection scope of the claims of the present invention.
Claims (7)
1. A centrifugal microfluidic detection system for seminal plasma fructose comprises a centrifuge and is characterized in that a centrifugal seminal plasma fructose analysis microfluidic chip is mounted on a main shaft of the centrifuge and comprises a central reagent cavity packaged with a pre-packaged reagent, a sample injection cavity packaged with alkaline solids, a separation quantitative cavity, a reagent quantitative cavity, a mixing cavity packaged with indole freeze-dried spheres and at least one test cavity packaged with protective agent freeze-dried spheres; wherein, central reagent chamber pass through the passageway with reagent ration chamber intercommunication, advance the appearance chamber pass through the passageway with separation ration chamber intercommunication, separation ration chamber with reagent ration chamber respectively pass through the passageway with the hybrid chamber intercommunication, the hybrid chamber pass through the passageway with test chamber intercommunication.
2. The centrifugal microfluidic seminal plasma fructose detection system of claim 1 wherein the sample waste chamber is in communication with the separation quantification chamber through a channel.
3. The centrifugal microfluidic seminal plasma fructose detection system of claim 1 wherein the reagent waste chamber is in communication with the reagent dosing chamber through a channel.
4. The centrifugal microfluidic detection system for fructose syrup of claim 1, wherein the mixed liquid waste chamber is communicated with the test chamber through a channel.
5. The centrifugal microfluidic seminal plasma fructose detection system of any one of claims 1-4 wherein the pre-packaged reagent is concentrated hydrochloric acid.
6. The centrifugal microfluidic seminal plasma fructose detection system of any one of claims 1-4 wherein the alkaline solids are sodium hydroxide solids.
7. A centrifugal microfluidic detection method for seminal plasma fructose is characterized by comprising the following steps:
s1, mixing the seminal plasma, the diluent and the deproteinized liquid, injecting the mixture into a sample injection cavity, and fully dissolving and reacting the mixture with alkaline solids to form a sample;
s2, starting a centrifugal machine to rotate the centrifugal type seminal plasma fructose analysis microfluidic chip, enabling a pre-packaged reagent in the central reagent cavity to enter a reagent quantitative cavity, and enabling a sample in the sample injection cavity to enter a separation quantitative cavity;
s3, starting the centrifugal machine to rotate the centrifugal seminal plasma fructose analysis microfluidic chip, enabling the sample in the separation quantitative cavity and the prepackaged reagent in the reagent quantitative cavity to respectively enter a mixing cavity, and dissolving the indole freeze-drying balls in the mixing cavity to form a mixed solution;
and S4, starting the centrifugal machine to rotate the centrifugal seminal plasma fructose analysis microfluidic chip, enabling the mixed liquid to enter the test cavity and mixing the mixed liquid with the protective agent freeze-drying balls in the test cavity.
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