CN112354571A - Multidimensional microfluidic electrophoresis chip, detection device and detection method - Google Patents
Multidimensional microfluidic electrophoresis chip, detection device and detection method Download PDFInfo
- Publication number
- CN112354571A CN112354571A CN202010770908.6A CN202010770908A CN112354571A CN 112354571 A CN112354571 A CN 112354571A CN 202010770908 A CN202010770908 A CN 202010770908A CN 112354571 A CN112354571 A CN 112354571A
- Authority
- CN
- China
- Prior art keywords
- electrophoresis
- channel
- chip
- isoelectric focusing
- storage unit
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Images
Classifications
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
- B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
- B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
- B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
- B01L3/502753—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip characterised by bulk separation arrangements on lab-on-a-chip devices, e.g. for filtration or centrifugation
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N27/00—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means
- G01N27/26—Investigating or analysing materials by the use of electric, electrochemical, or magnetic means by investigating electrochemical variables; by using electrolysis or electrophoresis
- G01N27/416—Systems
- G01N27/447—Systems using electrophoresis
- G01N27/44704—Details; Accessories
- G01N27/44717—Arrangements for investigating the separated zones, e.g. localising zones
- G01N27/44721—Arrangements for investigating the separated zones, e.g. localising zones by optical means
- G01N27/44726—Arrangements for investigating the separated zones, e.g. localising zones by optical means using specific dyes, markers or binding molecules
Abstract
This application at first proposes to design a parallel micro-fluidic electrophoresis chip of multidimension multichannel, has the changeover portion, the changeover portion includes semicircle or oval structure that a channel width narrows gradually at least, and it can improve the not good problem of protein separation effect, obtains high-resolution protein separation fingerprint. Meanwhile, a detection system is provided, which comprises the multi-dimensional microfluidic electrophoresis chip and can be applied to analysis and identification of biological samples such as microorganisms and clinical blood samples. Further, the method comprises a detection method which uses the detection device provided by the application to obtain the high-resolution separation image and analyze and identify the high-resolution separation image by means of the image.
Description
Technical Field
The invention relates to the technical field of microorganism detection, and relates to a multi-dimensional microfluidic electrophoresis chip, and a detection device and a detection method applying the multi-dimensional microfluidic electrophoresis chip.
Background
In the existing technical field of microorganism detection, when microorganism classification detection methods and devices such as morphological characteristics, physiological and biochemical reaction characteristics and the like are adopted for microorganism detection, microorganism culture takes long time and operation is complicated; the problems of high technical difficulty, easy occurrence of false positive and high cost exist by using nucleic acid detection molecular biology technologies based on 16S amplicon sequencing, QPCR, gene chips, metagenome and macrotranscriptome and the like; the detection device based on the immune method on the protein level has higher cost, and the species which can be detected is limited by the available antibodies.
At the beginning of this century, NISHIMOTOH in patent literature (JP 2003114215A) discloses an electrophoresis chip for chemical/biological analysis, which uses a simple "cross-shaped" structure to realize a western blot. However, the above-mentioned patent techniques are still only one-dimensional gel electrophoresis techniques in fact, and there is a limitation in the separation and analysis capabilities in the face of cells, microorganisms, and other living bodies having complex protein components.
The subsequent development of electrophoretic microfluidic chips over time has focused mainly on the design of "two-dimensional" electrophoretic microfluidic chips in order to increase the separation capacity of electrophoresis. For example, elegans patent document (CN 1469123A) discloses a microfluidic chip for protein analysis and its application in protein analysis, wherein the microfluidic chip is designed to further add a basic unit comprising a plurality of channels on the basis of a "cross" structure. For another example, the chip disclosed in the patent document (CN 1779431A) of linderan comprises a second unit, which is an electrophoretic separation analysis after protein concentration. The sample introduction channel of the chip can be in a T-shaped structure. Other patent technologies related to overall structure and morphological structure adjustment of the channel further include CN2831115Y, CN1786988A, CN104148123A, CN105092677A, CN104297327A, US2014332382 a1, US 8728290B 1, and the like.
To the best of the knowledge of the inventors of the present application, non-patent document 1(Emrich C A, Medintz I L, Chu W K, et al. Microfibrous Two-Dimensional electro-phoresis Device for Differential Protein Expression [ J ] Analytical Chemistry,2007,79(19):7360 and 7366.) first mentions that the ultrafine channels are processed between Two-Dimensional channels to make connections, creating resistance to prevent diffusion and at the same time reducing the volume of diffusion. Although the ultra-fine channel design implemented in the above-mentioned documents is not uncommon in the field of electrophoresis technology, for example US2011220499 a1, JP 2005233944A. However, the design of the channel in two dimensions to improve the protein separation capability requires a comprehensive consideration of the isoelectric focusing channel characteristics and the gel electrophoresis channel characteristics, which is significantly different from the general design in the electrophoresis field. As such, many subsequent technologies of multi-dimensional electrophoresis chips are referred to by reference, for example, non-patent document 2(West J, Becker M, tombrick S, et al. micro Total Analysis Systems: last Achievements [ J ]. Analytical Chemistry,2008,80(12):4403-
However, in the actual electrophoretic separation experiment process, the design method in non-patent document 1 still has a situation that the protein separation effect is not good enough or the protein analysis effect is not obvious from the multidimensional microfluidic electrophoresis chip without adding the ultrafine channel. In view of the above, the present application first proposes to design a multi-dimensional microfluidic electrophoresis chip, which can improve the problem of poor protein separation effect. Meanwhile, a detection system is provided, which comprises the multi-dimensional microfluidic electrophoresis chip, and can be applied to analysis and identification of biological samples such as microorganisms, including but not limited to detection of biological samples such as microorganisms based on protein separation fluorescence images. Further, the method for detecting the single and mixed microorganisms comprises the step of using the detection device provided by the application to identify the single and mixed microorganisms by acquiring high-resolution separation images and utilizing an image analysis technology. Technical implementation of specific separation and analysis can be referred to non-patent technical documents 3 to 10 provided below:
non-patent document 3, Fengming Lin, Xiaocha Zhao, Jianche Wang, Shiyong Yu, Xuefei Lv, Yulin Deng, Lina Geng, HuaJun Li, A novel microfluidic chip electrophoresis apparatus for laboratory, label-free, multi-protein detection based on a graphene energy transfer biosensor, analysis, 2014(139),2890 + 2895.
Non-patent technical literature 4, Lin Xia, FengMing Lin, Xin Wu, Jianshe Wang, Chuanli Liu, Qi Tang, Shiyong Yu, Kunjie Huang, XuefeiLv, Yulin ding, Lin gen g, On-chip protein isoelectric focusing using a photo-immobilized pH gradient, j.sep.sci.,2014 Nov; 37(21):3174-80
Non-patent document 5, Ni Hou, Yu Chen, Shiyong Yu, Zongliang Quan, Han Zhu, Chenhua Pan, Yulin Deng, Lina Geng, Native Protein Separation by Isoelectric Focusing and Blue Gel Electrophoresis-Coupled Two-dimensional Microfluidic Chip Electrophoresis, Chromatographania, 2014 77, 1339-
Non-patent document 6, Fengmin Lin, Shiyong Yu, Le Gu, Xuetao Zhu, Jiangshe Wang, Han Zhu, Yi Lu, Yihua Wang, Yulin Deng, Lina Geng, In situ photo-immobilized pH gradient electrochemical focusing and zone electrophoresis integrated two-dimensional microfluidic chip electrophoresis for protein separation, Microchip Acta,2015,182(13-14):2321-2328
Non-patent document 7, Shiyong Yu, Jiandong Xu, Kunjie Huang, Juan Chen, Jinyan Duan, Yuanqin Xu, Hong Qing, Lina Geng and Yulin Deng, alpha novel method to prediction protein aggregation using two-dimensional native protein microfluidic chip electrophoresis, anal. methods,2016,8,8306-
Non-patent technical literature 8, Yi Lu, Shiyong Yu, Fengming Lin, Fankai Lin, Xiaocha Zhuo, Liqing Wu, Yunfei Miao, Huangjun Li, Yulin Deng, Lina Geng, Simultaneous latex-free screening of G-quadruplex active library from natural media via a microfluidic chip architecture-based transducer array, analysis.2017,142 (22): 4257-.
Non-patent document 9, Zerong Liao, Jianfeng Wang, Pengjie Zhang, Yang Zhang, Yunfei Miao, Shimeng Gao, Yulin Deng, Lina Geng, Recent advances in microfluidic chip integrated electronic biosensions for multiplexed detection, Biosensors and Bioelectronics,2018,121(15)272-
Non-patent document 10, Zerong Liao, Yang Zhang, Yonggui Li, Yunfei Miao, Shimeng Gao, Yulin Deng, Lina Geng, Microfluidic chip coupled with optical biosensists for multiple analytes, A review, Biosensors and Bioelectronics, 2019,126(1)697-
Disclosure of Invention
In order to achieve the purpose, the invention adopts the following specific scheme:
the invention relates to a multidimensional microfluidic electrophoresis chip, which is provided with an electrophoresis carrying platform of an electrophoresis separation channel and comprises at least one group of buffer solution storage units (B, BW); at least comprises a group of loading/acid-alkali liquid storage units (S, SW); wherein the sample loading/acid-alkali liquid storage unit S and the SW are communicated through a chip electrophoresis channel; each buffer solution storage unit is communicated with the isoelectric focusing channel through a gel electrophoresis channel; at least part of the junction of the gel electrophoresis channel and the isoelectric focusing channel has a transition.
As just one embodiment of the invention, in particular, the transition section has a non-uniform cross-sectional area. The unevenness of the cross-sectional area is caused by unevenness of the transition section in the width direction, or the unevenness of the cross-sectional area is caused by unevenness of the transition section in the depth direction. Preferably, the transition section at least comprises a semicircular or elliptical structure with gradually narrowed channel width; preferably, the transition section has at least one series of symmetrical channel configurations.
As another embodiment of the invention only, in particular, the transition section comprises at least one asymmetric channel configuration. Preferably, the asymmetric channel shape may be an inverted trapezoidal structure with gradually narrowing channel width, or other common asymmetric geometric structures.
Specifically, the isoelectric focusing channel width is greater than the gel electrophoresis channel width.
Specifically, the isoelectric focusing channel is arc-shaped, the buffer solution storage unit B is an arc-shaped groove, and the buffer solution storage unit BW is located at the center of the isoelectric focusing channel.
Specifically, the isoelectric focusing channel is respectively connected with the buffer solution storage unit and the loading/acid-alkali solution storage unit.
Specifically, the buffer solution storage unit BW is arranged outside the electrophoresis carrier, the multidimensional microfluidic electrophoresis chip (3) further comprises a capillary, and the external buffer solution storage unit BW is connected with a gel electrophoresis channel arranged on the electrophoresis carrier through more than two capillaries.
The invention further relates to a detection device, which comprises the multi-dimensional microfluidic electrophoresis chip specifically defined by the invention, and further comprises a data acquisition module (1), a data analysis module (2) and a high-voltage power supply module (4), wherein the data acquisition module (1) is used for acquiring a separation image of an object to be detected; the data analysis module (2) is used for carrying out image processing on the separated images acquired by the data acquisition module (1) so as to obtain the component information of the object to be detected.
The invention further relates to a detection method, which comprises the detection device of the invention, and the electrophoresis and detection processes comprise the following steps:
step 1, washing and pretreating an electrophoresis channel of a multi-dimensional microfluidic electrophoresis chip;
step 2, injecting gel into an electrophoresis channel of the multidimensional microfluidic electrophoresis chip, and then applying voltage to the buffer solution storage unit B and the buffer solution storage unit BW for a period of time to complete pre-electrophoresis;
step 3, injecting a mixture of the sample and isoelectric focusing amphoteric electrolyte into an isoelectric focusing channel of the multidimensional microfluidic electrophoresis chip, or injecting the sample into an isoelectric focusing channel which is pre-formed with immobilized isoelectric focusing gradient;
step 4, applying voltage to the sample loading/acid-alkali liquid storage units S and SW at the two ends of the isoelectric focusing channel to perform isoelectric focusing, and stopping power supply after the isoelectric focusing is completed;
step 5, applying voltage at two ends of the gel electrophoresis channel to transfer the sample after the first-dimension isoelectric focusing is finished to a second-dimension gel electrophoresis channel, and then carrying out gel electrophoresis separation;
and 6, acquiring an electrophoresis separation image of the chip by using the data acquisition module, and analyzing the biological sample.
Has the advantages that:
the invention adopts the multidimensional microfluidic electrophoresis chip to carry out electrophoretic separation, the multidimensional microfluidic electrophoresis chip is provided with the isoelectric focusing channel and more than two gel electrophoresis channels, and the more than two gel electrophoresis channels improve the separation flux of gel electrophoresis in the lower gel electrophoresis channel, thereby obtaining a high-resolution separation fluorescence image. Then, the rapid identification of the microorganisms is realized by means of an image data analysis tool and means, and the device has low cost and high identification efficiency.
In the multidimensional microfluidic electrophoresis chip, the non-uniform cross-sectional area design of the transition section can further improve fluid flow and separation, so that a separation map with higher resolution is obtained.
For some embodiments, the transition section is modified by adjusting the channel width and the structural configuration. Particularly, a circular or elliptical structure is adopted, so that the liquid storage capacity is higher on the premise of a given length and/or depth, mainly because the circular structure is a geometrical structure with the largest area under the condition of a given perimeter. According to the technical scheme, the aggregation liquid storage capacity can be improved by at least 20 percent compared with that of non-patent technical document 1 by adjusting the local form.
For some embodiments, the transition section is improved by having an asymmetric channel structure, and the asymmetric structure can cause inconsistent resistance on two sides of the channel structure, so that the speed of protein migration on one side is different from that on the other side, thereby being more beneficial to realize the sequential separation and migration of proteins in the gel electrophoresis channel and further obtaining a separation map with higher resolution (more protein spectral peaks).
For some embodiments, the transition is improved by having at least one set of symmetrical channel configurations in series, using multiple band accumulation effects, reducing band spreading, and increasing detection sensitivity.
Drawings
FIG. 1 is an overall device diagram of the present invention, wherein 1-data acquisition module, 2-data analysis module, 3-multidimensional microfluidic electrophoresis chip, 4-high voltage power module;
FIG. 2 is a schematic diagram of a channel structure of a common array type multi-dimensional microfluidic electrophoresis chip;
FIG. 3 is a diagram of a sample simulation of a common array type multi-dimensional microfluidic electrophoresis chip
FIG. 4 is a schematic diagram of a divergent array channel type multi-dimensional microfluidic electrophoresis chip channel structure;
FIG. 5 is a schematic diagram of a channel structure of a multidimensional microfluidic electrophoresis chip externally connected with an array capillary;
FIG. 6 is a schematic diagram of a multi-dimensional microfluidic electrophoresis chip channel structure of coupled plate gel electrophoresis;
FIG. 7 is a schematic view of a different transition section configuration;
FIG. 8 is a schematic diagram of a channel structure of a multi-dimensional microfluidic electrophoresis chip with a buffer solution storage unit in a separated mode;
FIG. 9 is a schematic diagram of a channel structure of a multi-dimensional microfluidic electrophoresis chip with a segmented buffer solution storage unit;
FIG. 10 is a schematic diagram of the channel structure of the multidimensional microfluidic electrophoresis chip with the sample loading liquid storage units (S and SW) separated from the acid-base liquid storage units (A and B).
Detailed Description
The invention is described in detail below by way of example with reference to the accompanying drawings.
The micro-fluidic chip is the leading field of the development of modern biological analysis devices to miniaturization, integration and automation, and is expected to provide a quick, simple and efficient platform for detecting and quantifying molecules of microorganisms. The multidimensional microfluidic electrophoresis chip has the advantage of easy integration, and can obtain higher separation capacity by constructing a multidimensional separation platform, thereby creating opportunities for obtaining high-resolution separation images and realizing identification/detection of biological samples such as microorganisms.
Example 1: the invention provides a microorganism detection device based on a multidimensional microfluidic electrophoresis chip, which is integrally arranged as shown in figure 1 and comprises a multidimensional microfluidic electrophoresis chip 3, a data acquisition module 1, a data analysis module 2 and a high-voltage power supply module 4;
the multi-dimensional microfluidic electrophoresis chip 3 is used for generating a separation map of a sample;
the multi-dimensional microfluidic electrophoresis chip 3 comprises at least four liquid storage units and an electrophoresis carrying platform provided with an electrophoresis separation channel;
the high-voltage power supply module 4 is used for providing an electrophoresis power supply for the multi-dimensional microfluidic electrophoresis chip, outputting multi-path voltage of 0-5000V, and is controlled by programmable multi-step voltage, fast in switching and stable in output voltage.
The four liquid storage units are respectively marked as a buffer liquid storage unit B, a buffer liquid storage unit BW, a sample loading/acid-alkali liquid storage unit S and a sample loading/acid-alkali liquid storage unit SW; the electrophoresis liquid comprises liquids used in all stages of electrophoresis;
the electrophoresis separation channel comprises a gel electrophoresis channel and an isoelectric focusing channel, and the gel electrophoresis channel comprises an upper gel electrophoresis channel and a lower gel electrophoresis channel;
the sample loading/acid-alkali liquid storage unit S and the sample loading/acid-alkali liquid storage unit SW are communicated through a channel, and the channel is an isoelectric focusing channel; the buffer solution storage unit B and the buffer solution storage unit BW are respectively arranged on two sides of the isoelectric focusing channel; the buffer solution storage unit is communicated with the isoelectric focusing channel through at least two channels, namely a channel between the buffer solution storage unit B and the isoelectric focusing channel is an upper gel electrophoresis channel, and a channel between the buffer solution storage unit BW and the isoelectric focusing channel is a lower gel electrophoresis channel;
the data acquisition module 1 is used for acquiring a separation image obtained by electrophoretic separation of the multidimensional microfluidic electrophoresis chip 3, the data acquisition is realized based on optical detection, the separation image in the fluorescence inverted microscope can be acquired by an inverted microscope and aiming at the end region of an array channel or an array capillary, and the data acquisition can also be realized by installing an array optical fiber at the end of the array channel or the capillary.
The data acquisition of multiple channels can be carried out by adopting image acquisition devices such as an inverted microscope and the like, and the separation signals of each channel can be respectively collected by adopting array optical fibers and the like.
The data analysis module 2 is used for processing the images acquired by the data acquisition module 1 to obtain the component information of the microorganism to be detected.
The image processing step includes:
reading in a separation image;
preprocessing the separated image, mainly including image normalization and simple noise removal;
analyzing the image data histogram of the separated image, and calculating the entropy value of the image and the statistic of the corresponding distribution, wherein the reference indexes are kurtosis and skewness respectively, and obtaining the component information of the microorganism to be detected.
In order to facilitate sample loading, the width of the isoelectric focusing channel is larger than that of the gel electrophoresis channel; the multi-dimensional microfluidic electrophoresis chip is a basic form of coupling isoelectric focusing and gel electrophoresis, and the structural design of the multi-dimensional microfluidic electrophoresis chip aims to improve the separation flux of gel electrophoresis in a gel electrophoresis channel so as to obtain a high-resolution separation fluorescence image. A first-dimension isoelectric focusing channel and a second-dimension gel electrophoresis channel are arranged in the multi-dimensional microfluidic electrophoresis chip, and the second-dimension gel electrophoresis channel adopts a multi-channel parallel structure, so that the separation flux of chip electrophoresis is improved.
The electrophoresis separation channel of the multi-dimensional microfluidic electrophoresis chip shown in fig. 2 is a common array type multi-dimensional microfluidic electrophoresis chip channel structure, an isoelectric focusing channel is arranged between a sample loading/acid-alkali liquid storage unit S and a sample loading/acid-alkali liquid storage unit SW in the left picture of fig. 2, and the length, the width and the depth of the isoelectric focusing channel are 23mm, 150um and 30um respectively; the isoelectric focusing channels are linear, the buffer solution storage unit B and the buffer solution storage unit BW are linear grooves, the gel electrophoresis channels are parallel to each other, 16 parallel upper gel electrophoresis channels are arranged between the isoelectric focusing channels and the buffer solution storage unit B, and the conventional length, width and depth of the upper gel electrophoresis channels are respectively 18mm, 50um and 30 um; between the isoelectric focusing channel and the liquid storage unit BW are 16 parallel lower gel electrophoresis channels, the conventional length, width and depth of which are 32mm, 100um and 30um respectively.
The multi-dimensional microfluidic electrophoresis chip is manufactured through the steps of mask manufacturing, photolithography, wet etching, punching, bonding and the like, and the obtained entity is shown in fig. 3, and the manufacturing process steps can be specifically referred to non-patent technical document 11 (manufacturing method of protein separation glass microfluidic chip, gunna, all-purpose, hounie, chenyan, hanna, xujia, hudingyu, li, dunlin, university of beijing physics, 2013,33(4), 436-.
Example 2: a divergent array channel type multidimensional microfluidic electrophoresis chip, which is different from that of embodiment 1 mainly in that the electrophoretic separation channel structure of the multidimensional microfluidic electrophoresis chip is different, the channel structure of the divergent array channel type multidimensional microfluidic electrophoresis chip is shown in fig. 4, an isoelectric focusing channel from a sample loading/acid-alkali solution storage unit S to a sample loading/acid-alkali solution storage unit SW is arc-shaped, a buffer solution storage unit B is arc-shaped grooves, the upper gel electrophoresis channels are parallel to each other, a buffer solution storage unit BW is positioned at the center of the isoelectric focusing channel, and the buffer solution storage unit BW can adopt arc-shaped grooves or circular grooves; when the buffer solution storage unit BW adopts the arc-shaped groove, more than two lower gel electrophoresis channels of the buffer solution storage unit BW are divergently connected to the isoelectric focusing channel; when the buffer solution storage unit BW adopts a circular groove, the lower gel electrophoresis channel of the buffer solution storage unit BW is divergently connected to the isoelectric focusing channel.
Example 3: the microfluidic chip of external array capillary, the electrophoresis microscope carrier can adopt the same structure as embodiment 1 or embodiment 2, but buffer solution stock solution unit BW sets up outside the electrophoresis microscope carrier, in addition, multidimensional microfluidic electrophoresis chip still includes the capillary, as shown in figure 5, external buffer solution stock solution unit BW links to each other with the lower gel electrophoresis passageway of setting on the electrophoresis microscope carrier through two or more capillaries, lower gel electrophoresis passageway passes through capillary and buffer solution stock solution unit BW intercommunication.
Example 4: as shown in fig. 6, a lower gel electrophoresis channel of the microfluidic chip for coupled plate gel electrophoresis includes two or more connection channels and gel electrophoresis grooves, the connection channels are respectively connected with the gel electrophoresis grooves, and then are connected with a buffer solution storage unit BW through the gel electrophoresis grooves, and other parts are the same as those in embodiment 1 or embodiment 2.
Example 5: the microfluidic chip is provided with a transition section, the transition section is arranged on a channel section at the joint of the isoelectric focusing channel and the gel electrophoresis channel, and the transition section can be specifically arranged as shown in a reference figure 7. The rest of the structure may be any one of the embodiments as in embodiments 1 to 4, and enlarged views of the inverted funnel-shaped structure of non-patent document 1 are shown in fig. 2 to 6. The gel electrophoresis channel starts with a narrow width and then widens, starting from the isoelectric focusing channel, which is shown in the figure as an inverted funnel structure in a partially enlarged view. The improved structure of the transition section can further effectively reduce the concentration diffusion of the sample after sample loading, reduce the suction force to the cellulose gel in the gel electrophoresis channel when the negative pressure is applied to the liquid storage unit connected with the isoelectric focusing channel, ensure the smooth cleaning process of the gel electrophoresis channel during sample loading, improve the separation flux of the gel electrophoresis of the sample during electrophoretic separation, and contribute to obtaining a high-resolution separation image.
Taking a protein sample as an example, the process of performing electrophoretic separation on the sample based on the multidimensional microfluidic electrophoresis chip comprises the following steps:
a. chip pretreatment: the newly prepared chip is treated by 1mol/L hydrochloric acid for 1 hour and then treated by triple distilled water for 0.5 hour. Before each experiment, the sample is washed by triple distilled water for 10min, treated by 1mol/L NaOH solution for 10min, washed by triple distilled water for 10min, washed by BSA (or other coating substances) dissolved by PBS for 30min, and treated by methylcellulose solution added with BSA for 15 min. After each experiment, the mixture was washed with triple distilled water for 10min and then with 98% H2SO4And processing the channel.
b. Loading:
after washing the chip channels, respectively sucking residual liquid of 4 liquid storage units, wiping the surface of the chip by using filter paper, and then injecting BSA cellulose gel with the same volume as that of the liquid storage units;
secondly, electrifying two ends of the buffer solution storage unit B and the buffer solution storage unit BW for a period of time to finish pre-electrophoresis;
respectively sucking residual liquid of 4 liquid storage units, injecting ultrapure water with the same volume as the sample loading/acid-alkali liquid storage unit S, and simultaneously injecting BSA cellulose gel with the same volume as the buffer liquid storage unit B and the buffer liquid storage unit BW into the two liquid storage units;
fourthly, adding larger negative pressure to the sample loading/acid-alkali liquid storage unit SW until the liquid level of the triple distilled water in the sample loading/acid-alkali liquid storage unit S can be quickly reduced;
keeping the buffer solution storage unit B and the buffer solution storage unit BW unchanged, and respectively sucking residual liquid of the sample loading/acid alkali solution storage unit S and residual liquid of the sample loading/acid alkali solution storage unit SW after the sample loading/acid alkali solution storage unit S and the sample loading/acid alkali solution storage unit SW are switched;
sixthly, injecting the sample into a sample loading/acid-alkali liquid storage unit S, and then applying short-term and extremely-small negative pressure to the sample loading/acid-alkali liquid storage unit SW; making the sample enter a one-dimensional separation channel, namely an isoelectric focusing channel;
and seventhly, absorbing the residual liquid of the sample loading/acid-alkali liquid storage unit S and the residual liquid of the sample loading/acid-alkali liquid storage unit SW, adding acid and alkali liquid with the same volume respectively, and then compensating the liquid levels of the buffer liquid storage unit B and the buffer liquid storage unit BW.
c. Electrophoretic separation: firstly, an isoelectric focusing experiment is carried out, wherein the anode and the cathode of a power supply are respectively added to an acid-base pool, and voltage is applied to carry out isoelectric focusing. And after focusing is finished, removing the isoelectric focusing electrode, then carrying out a gel electrophoresis experiment, adding a positive electrode into a buffer solution storage unit B, adding a negative electrode into a buffer solution storage unit BW, and adjusting the position of the chip in real time according to the position of the separation strip to ensure that the separation strip is always positioned within the visual field of the fluorescence induction microscope. In the experiment, the multidimensional microfluidic electrophoresis chip disclosed by the invention is used for separating escherichia coli holoprotein, bacillus subtilis holoprotein and staphylococcus aureus holoprotein.
After the protein is primarily separated, a data acquisition module is used for intercepting one picture every 2min, 15 pictures are intercepted in total, noise removal is carried out on the pictures, primary processing of gray level image conversion is carried out, a method based on peak intensity probability distribution is adopted for feature extraction, fingerprint features of a spectrogram are formed, and then a multivariate statistical technology is adopted for reducing the dimension of the multidimensional features to two dimensions. And (4) testing the unknown sample after finishing classification training according to the known microorganism type information by using the training set sample.
Example 6: the buffer solution storage unit is a separated multidimensional microfluidic electrophoresis chip, as shown in figure 8. The difference from the divergent array channel type multidimensional microfluidic electrophoresis chip in the embodiment 2 is mainly that the buffer liquid pool adopts an isolated type, each liquid pool is provided with one electrode, and a plurality of electrodes are connected in parallel.
Example 7: the buffer solution storage unit is a segmented multidimensional microfluidic electrophoresis chip as shown in figure 9. The difference from the divergent array channel type multi-dimensional microfluidic electrophoresis chip in the embodiment 2 is mainly that the length of the buffer solution pool is reduced, each solution pool is provided with one electrode, and the multiple electrodes are connected in parallel.
Example 8: FIG. 10 shows a multi-dimensional microfluidic electrophoresis chip with a sample loading reservoir unit (S and SW) and an acid-base solution reservoir unit (A and B) separated. The difference from the divergent array channel type multi-dimensional microfluidic electrophoresis chip in example 2 is that an additional set of acid-base cells is separately arranged and separated from the sample cells S and SW. When the chip shown in example 10 was subjected to electrophoresis, after completion of the sample application between the sample cells S and SW, the sample was directly applied to the pH cell to perform isoelectric focusing, and then applied to the B and BW cells to perform the second dimension gel electrophoresis.
It should be noted that the use of "including," "comprising," or "having" and variations thereof herein is meant to encompass the items listed thereafter and equivalents thereof as well as additional items. Unless otherwise limited, the terms "interface," "communication," and variations thereof are used broadly herein and include direct and indirect communication, interfacing.
The dimensions and values disclosed herein are not to be understood as being strictly limited to the exact numerical values recited. Rather, unless otherwise specified, each such dimension is intended to mean both the recited value and a functionally equivalent range surrounding that value. For example, a dimension disclosed as "40 mm" is intended to mean "about 40 mm".
All documents cited in the text of the invention are incorporated by reference herein in relevant part. The citation of any document is not to be construed as an admission that it is prior art with respect to the present invention. To the extent that any meaning or definition of a term in this document conflicts with any meaning or definition of the same term in a document incorporated by reference, the meaning or definition assigned to that term in this document shall govern.
The principles, representative embodiments and modes of operation of the present invention have been described in the foregoing description. However, the several aspects of the invention that are intended to be protected are not to be construed as limited to the particular embodiments disclosed. Furthermore, the embodiments described herein are to be considered as illustrative and not restrictive. It is to be understood that changes and variations may be made by use of other and equivalent concepts without departing from the spirit of the invention. Accordingly, it is expressly intended that all such variations, changes and equivalents fall within the spirit and scope of the claimed invention.
Claims (7)
1. The utility model provides a multidimension micro-fluidic electrophoresis chip, multidimension micro-fluidic electrophoresis chip is equipped with the electrophoresis microscope carrier of electrophoresis separation channel, its characterized in that:
comprising at least one set of buffer reservoir units (B, BW);
at least comprises a group of loading/acid-alkali liquid storage units (S, SW);
wherein the sample loading/acid-alkali liquid storage unit S and the SW are communicated through a chip electrophoresis channel;
each buffer solution storage unit is communicated with the isoelectric focusing channel through a gel electrophoresis channel; at least part of the junction of the gel electrophoresis channel and the isoelectric focusing channel has a transition.
2. A multi-dimensional microfluidic electrophoresis chip according to claim 1 or 2 wherein said transition section comprises at least one semicircular or elliptical structure with a gradually narrowing channel width.
3. The multi-dimensional microfluidic electrophoresis chip of claim 1 wherein said isoelectric focusing channel width is greater than said gel electrophoresis channel width.
4. The multi-dimensional microfluidic electrophoresis chip of claim 1 wherein said isoelectric focusing channels are arc-shaped, said buffer reservoir unit B is an arc-shaped groove, and said buffer reservoir unit BW is located at the center of the isoelectric focusing channels.
5. The multi-dimensional microfluidic electrophoresis chip according to claim 1, wherein the buffer solution storage unit BW is specifically disposed outside the electrophoresis carrier, the multi-dimensional microfluidic electrophoresis chip (3) further comprises a capillary, and the external buffer solution storage unit BW is connected to the gel electrophoresis channel disposed on the electrophoresis carrier through two or more capillaries.
6. A detection device comprising the multi-dimensional microfluidic electrophoresis chip defined in claims 1 to 5, respectively, and further comprising a data acquisition module (1), a data analysis module (2), and a high voltage power supply module (4), the data acquisition module (1) being configured to acquire a separation image of an object to be detected;
the data analysis module (2) is used for carrying out image processing on the separated images acquired by the data acquisition module (1) so as to obtain the component information of the object to be detected.
7. A detection method comprising the detection device of claim 6, the electrophoresis and detection process comprising the steps of:
step 1, washing and pretreating an electrophoresis channel of a multi-dimensional microfluidic electrophoresis chip;
step 2, injecting gel into an electrophoresis channel of the multidimensional microfluidic electrophoresis chip, and then applying voltage to the buffer solution storage unit B and the buffer solution storage unit BW for a period of time to complete pre-electrophoresis;
step 3, injecting a mixture of the sample and isoelectric focusing amphoteric electrolyte into an isoelectric focusing channel of the multidimensional microfluidic electrophoresis chip, or injecting the sample into an isoelectric focusing channel which is pre-formed with immobilized isoelectric focusing gradient;
step 4, applying voltage to the sample loading/acid-alkali liquid storage units S and SW at the two ends of the isoelectric focusing channel to perform isoelectric focusing, and stopping power supply after the isoelectric focusing is completed;
step 5, applying voltage at two ends of the gel electrophoresis channel to transfer the sample after the first-dimension isoelectric focusing is finished to a second-dimension gel electrophoresis channel, and then carrying out gel electrophoresis separation;
and 6, acquiring an electrophoresis separation image of the chip by using the data acquisition module, and analyzing the biological sample.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010770908.6A CN112354571B (en) | 2019-07-11 | 2019-07-11 | Multidimensional microfluidic electrophoresis chip, detection device and detection method |
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202010770908.6A CN112354571B (en) | 2019-07-11 | 2019-07-11 | Multidimensional microfluidic electrophoresis chip, detection device and detection method |
| CN201910622879.6A CN110327994B (en) | 2019-07-11 | 2019-07-11 | Multidimensional microfluidic electrophoresis chip, detection device and detection method |
Related Parent Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201910622879.6A Division CN110327994B (en) | 2019-07-11 | 2019-07-11 | Multidimensional microfluidic electrophoresis chip, detection device and detection method |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN112354571A true CN112354571A (en) | 2021-02-12 |
| CN112354571B CN112354571B (en) | 2022-02-11 |
Family
ID=68146326
Family Applications (4)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202010770908.6A Active CN112354571B (en) | 2019-07-11 | 2019-07-11 | Multidimensional microfluidic electrophoresis chip, detection device and detection method |
| CN202010771073.6A Active CN112354572B (en) | 2019-07-11 | 2019-07-11 | Multidimensional microfluidic electrophoresis chip, detection device and detection method |
| CN201910622879.6A Active CN110327994B (en) | 2019-07-11 | 2019-07-11 | Multidimensional microfluidic electrophoresis chip, detection device and detection method |
| CN202010770903.3A Active CN112354570B (en) | 2019-07-11 | 2019-07-11 | Multidimensional microfluidic electrophoresis chip, detection device and detection method |
Family Applications After (3)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202010771073.6A Active CN112354572B (en) | 2019-07-11 | 2019-07-11 | Multidimensional microfluidic electrophoresis chip, detection device and detection method |
| CN201910622879.6A Active CN110327994B (en) | 2019-07-11 | 2019-07-11 | Multidimensional microfluidic electrophoresis chip, detection device and detection method |
| CN202010770903.3A Active CN112354570B (en) | 2019-07-11 | 2019-07-11 | Multidimensional microfluidic electrophoresis chip, detection device and detection method |
Country Status (1)
| Country | Link |
|---|---|
| CN (4) | CN112354571B (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111250182B (en) * | 2020-02-11 | 2021-03-19 | 北京理工大学 | High-flux microfluidic electrophoresis screening chip and preparation method and application method thereof |
Citations (13)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1993005150A1 (en) * | 1991-09-05 | 1993-03-18 | Schering Aktiengesellschaft | Collagen-induced platelet aggregation inhibitor |
| CN1477391A (en) * | 2002-08-21 | 2004-02-25 | 中国科学院大连化学物理研究所 | Method for sedparating bialogical macromolecule by using two-dimensinal or multi-dimensinal capillary electrophoresis and its used interface |
| CN1493001A (en) * | 2001-03-16 | 2004-04-28 | ʩ��ķ����ϵͳ��˾ | Array-based biomolecule analysis |
| US20060068373A1 (en) * | 2004-09-24 | 2006-03-30 | Arijit Bose | Systems and methods of identifying biomarkers for subsequent screening and monitoring of diseases |
| JP2007064848A (en) * | 2005-08-31 | 2007-03-15 | Sharp Corp | Automated two-dimensional electrophoretic device, and device component |
| US20110120867A1 (en) * | 2007-08-22 | 2011-05-26 | Aida Engineering , Ltd. | Micro-channel chip for electrophoresis and method for electrophoresis |
| CN102183388A (en) * | 2011-01-25 | 2011-09-14 | 北京理工大学 | Preparation method of free flow electrophoresis chip with two-dimensional depth |
| CN102822672A (en) * | 2010-01-08 | 2012-12-12 | 安科健康公司 | High throughput cell-based HPV immunoassays for diagnosis and screening of HPV-associated cancers |
| CN204346798U (en) * | 2014-12-30 | 2015-05-20 | 北京理工大学 | A kind of protein nucleic acid tripping device |
| US20150260682A1 (en) * | 2014-03-14 | 2015-09-17 | Agency For Science, Technology And Research | Microfluidic device for gel electrophoresis and method of manufacturing thereof |
| CN106999797A (en) * | 2014-12-05 | 2017-08-01 | 先进电泳方案有限公司 | Apparatus and method for separating molecule |
| CN109750116A (en) * | 2019-03-12 | 2019-05-14 | 西安医学院 | PCR- internal control nucleic acid test strip detection method for mycoplasma pneumoniae detection |
| US10329547B1 (en) * | 2018-06-13 | 2019-06-25 | Caribou Biosciences, Inc. | Engineered cascade components and cascade complexes |
Family Cites Families (20)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| AU4323602A (en) * | 2000-11-20 | 2002-06-24 | 20 20 Gene Systems Inc | Methods, devices, arrays and kits for detecting and analyzing biomolecules |
| WO2003008977A2 (en) * | 2001-07-16 | 2003-01-30 | Protein, Forest, Inc. | Arrays of buffers for analysing biomolecules by their isoelectric point |
| EP1525464A2 (en) * | 2002-06-18 | 2005-04-27 | Invitrogen Corporation | Methods and apparatus for low resistance electrophoresis of prior-cast, hydratable separation media |
| US7699767B2 (en) * | 2002-07-31 | 2010-04-20 | Arryx, Inc. | Multiple laminar flow-based particle and cellular separation with laser steering |
| CN1287145C (en) * | 2003-07-14 | 2006-11-29 | 浙江大学 | Analysis method for micro flow control chip unicell |
| EP1612550B1 (en) * | 2004-07-03 | 2007-04-11 | Roche Diagnostics GmbH | Preconcentration interface coupling liquid chomatography to capillary electrophoresis |
| WO2007022026A2 (en) * | 2005-08-11 | 2007-02-22 | Biotrove, Inc. | Apparatus for assay, synthesis and storage, and methods of manufacture, use, and manipulation thereof |
| EP1804058A1 (en) * | 2005-12-28 | 2007-07-04 | Roche Diagnostics GmbH | Integrated two-dimensional gel electrophoresis |
| WO2010048173A2 (en) * | 2008-10-20 | 2010-04-29 | Stc.Unm | High resolution focusing and separation of proteins in nanofluidic channels |
| US20110244500A1 (en) * | 2010-04-01 | 2011-10-06 | Shapiro Howard K | Gel electrophoresis method useful for resolution and characterization of nerve tissue ultra high molecular weight protein aggregates |
| US10078075B2 (en) * | 2011-12-09 | 2018-09-18 | Vanderbilt University | Integrated organ-on-chip systems and applications of the same |
| CN102252878B (en) * | 2011-03-20 | 2013-02-13 | 北京理工大学 | Off-Gel free flow electrophoresis coupling chip and making method thereof |
| CN102788831B (en) * | 2012-08-13 | 2014-07-30 | 中国科学院研究生院 | Microfluidic chip electrophoretic-electrochemical detecting device with adjustable pH after separation and use thereof |
| GB201320146D0 (en) * | 2013-11-14 | 2014-01-01 | Cambridge Entpr Ltd | Fluidic separation and detection |
| JP6067655B2 (en) * | 2014-11-20 | 2017-01-25 | シャープ株式会社 | Transfer film with frame member, biomolecule analyzer, reagent tank and shaking device |
| CN105241984B (en) * | 2015-10-23 | 2018-10-23 | 厦门色谱分析仪器有限公司 | Array multidimensional liquid chromatogram column system |
| WO2017083609A1 (en) * | 2015-11-13 | 2017-05-18 | Life Technologies Corporation | Preparation of electrophoresis gels, and related devices, systems, and methods |
| JP7107842B6 (en) * | 2016-01-13 | 2023-08-18 | プロテインシンプル | system |
| CN105738637B (en) * | 2016-02-01 | 2017-09-29 | 北京理工大学 | The full-automatic capillary electrophoresis of analysis is separated suitable for space biological sample |
| CN107271685A (en) * | 2017-06-30 | 2017-10-20 | 中国农业大学 | A kind of method and its dedicated kit for detecting the content of fowl leukocyte interleukin 2 |
-
2019
- 2019-07-11 CN CN202010770908.6A patent/CN112354571B/en active Active
- 2019-07-11 CN CN202010771073.6A patent/CN112354572B/en active Active
- 2019-07-11 CN CN201910622879.6A patent/CN110327994B/en active Active
- 2019-07-11 CN CN202010770903.3A patent/CN112354570B/en active Active
Patent Citations (13)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1993005150A1 (en) * | 1991-09-05 | 1993-03-18 | Schering Aktiengesellschaft | Collagen-induced platelet aggregation inhibitor |
| CN1493001A (en) * | 2001-03-16 | 2004-04-28 | ʩ��ķ����ϵͳ��˾ | Array-based biomolecule analysis |
| CN1477391A (en) * | 2002-08-21 | 2004-02-25 | 中国科学院大连化学物理研究所 | Method for sedparating bialogical macromolecule by using two-dimensinal or multi-dimensinal capillary electrophoresis and its used interface |
| US20060068373A1 (en) * | 2004-09-24 | 2006-03-30 | Arijit Bose | Systems and methods of identifying biomarkers for subsequent screening and monitoring of diseases |
| JP2007064848A (en) * | 2005-08-31 | 2007-03-15 | Sharp Corp | Automated two-dimensional electrophoretic device, and device component |
| US20110120867A1 (en) * | 2007-08-22 | 2011-05-26 | Aida Engineering , Ltd. | Micro-channel chip for electrophoresis and method for electrophoresis |
| CN102822672A (en) * | 2010-01-08 | 2012-12-12 | 安科健康公司 | High throughput cell-based HPV immunoassays for diagnosis and screening of HPV-associated cancers |
| CN102183388A (en) * | 2011-01-25 | 2011-09-14 | 北京理工大学 | Preparation method of free flow electrophoresis chip with two-dimensional depth |
| US20150260682A1 (en) * | 2014-03-14 | 2015-09-17 | Agency For Science, Technology And Research | Microfluidic device for gel electrophoresis and method of manufacturing thereof |
| CN106999797A (en) * | 2014-12-05 | 2017-08-01 | 先进电泳方案有限公司 | Apparatus and method for separating molecule |
| CN204346798U (en) * | 2014-12-30 | 2015-05-20 | 北京理工大学 | A kind of protein nucleic acid tripping device |
| US10329547B1 (en) * | 2018-06-13 | 2019-06-25 | Caribou Biosciences, Inc. | Engineered cascade components and cascade complexes |
| CN109750116A (en) * | 2019-03-12 | 2019-05-14 | 西安医学院 | PCR- internal control nucleic acid test strip detection method for mycoplasma pneumoniae detection |
Non-Patent Citations (6)
| Title |
|---|
| BROMBERG, AVRAHAM等: "Microfabricated Linear Hydrogel Microarray for Single-Nucleotide Polymorphism Detection", 《ANALYTICAL CHEMISTRY》 * |
| EMRICH C A等: "Microfabricated Two-Dimensional Electrophoresis Device for Differential Protein Expression Profiling", 《ANALYTICAL CHEMISTRY》 * |
| IWAMOTO, M.等: "Antioxidant effects of carnitine supplementation on 14-3-3 protein isoforms in the aged rat hippocampus detected using fully automated two-dimensional chip gel electrophoresis", 《FREE RADICAL RESEARCH》 * |
| LIM, AMARETH等: "Characterization of a cathepsin D protease from CHO cell-free medium and mitigation of its impact on the stability of a recombinant therapeutic protein", 《BIOTECHNOLOGY PROGRESS》 * |
| 曲由等: "双向电泳关键因素分析", 《现代生物医学进展》 * |
| 许锦钢: "基于液滴微流控技术的二维液滴阵列生成系统的研究", 《中国优秀博硕士学位论文全文数据库(硕士) 工程科技Ⅰ辑 （月刊） 2019年》 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN112354572A (en) | 2021-02-12 |
| CN112354571B (en) | 2022-02-11 |
| CN110327994B (en) | 2020-12-08 |
| CN112354570B (en) | 2022-02-11 |
| CN112354570A (en) | 2021-02-12 |
| CN110327994A (en) | 2019-10-15 |
| CN112354572B (en) | 2022-02-11 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| US10232371B2 (en) | Microfluidic devices and methods for cell processing | |
| Kohlheyer et al. | Miniaturizing free‐flow electrophoresis–a critical review | |
| Hughes et al. | Microfluidic integration for automated targeted proteomic assays | |
| US20170160197A1 (en) | Method and device for single molecule imaging | |
| JP2003501639A (en) | Microfluidic devices for transverse and isoelectric focusing | |
| US20150099660A1 (en) | Multi-directional microfluidic devices comprising a pan-capture binding region and methods of using the same | |
| Gao et al. | Detection of urine protein by a paper-based analytical device enhanced with ion concentration polarization effect | |
| CN104646079B (en) | A kind of centrifugal microfluidic control chip for capillary electrophoresis and capillary gel electrophoresis device | |
| CN113227754A (en) | Image-based assays using smart monitoring structures | |
| CN112354571B (en) | Multidimensional microfluidic electrophoresis chip, detection device and detection method | |
| Hung et al. | A portable sample concentrator on paper-based microfluidic devices | |
| CA2969855A1 (en) | Apparatus and method for separating molecules | |
| US11376590B2 (en) | Diagnostic system for hemoglobin analysis | |
| Grist et al. | 3D projection electrophoresis for single-cell immunoblotting | |
| CN112239719A (en) | Digital PCR amplification device based on micropore array chip and method for amplifying by using same | |
| CN106846310A (en) | A kind of pathology aided analysis method based on immunohistochemistry technique | |
| CN110468027B (en) | Cell sorting microfluidic chip based on coaxial double-waveguide optical fiber | |
| CN111812091A (en) | Chip gel electrophoresis and on-line UV-VIS imaging detection device thereof | |
| KR102064388B1 (en) | Single point detection type microfluidic isoelectric focusing assay and chips using the same | |
| US20160018360A1 (en) | Microfluidic device for detecting nucleic acids and associated methods | |
| Quan et al. | Development of high-resolution multidimensional native protein microfluidic chip electrophoresis fingerprinting and its application in the quick analysis of unknown microorganisms | |
| Zhou et al. | Development and prospects of microfluidic platforms for sperm inspection | |
| Sami et al. | AQAFI: a bioanalytical method for automated KPIs quantification of fluorescent images of human leukocytes and micro–nano particles | |
| JP2004132952A (en) | Improvement in analytical method | |
| CN111172010B (en) | Cell sorting microfluidic chip based on three-core optical fiber |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |