CN110252435A - A kind of magnetic particle shines micro-fluidic chip and reaction method - Google Patents
A kind of magnetic particle shines micro-fluidic chip and reaction method Download PDFInfo
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- CN110252435A CN110252435A CN201910567512.9A CN201910567512A CN110252435A CN 110252435 A CN110252435 A CN 110252435A CN 201910567512 A CN201910567512 A CN 201910567512A CN 110252435 A CN110252435 A CN 110252435A
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- 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
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/5302—Apparatus specially adapted for immunological test procedures
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L2300/00—Additional constructional details
- B01L2300/08—Geometry, shape and general structure
- B01L2300/0809—Geometry, shape and general structure rectangular shaped
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L2300/00—Additional constructional details
- B01L2300/08—Geometry, shape and general structure
- B01L2300/0887—Laminated structure
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- Health & Medical Sciences (AREA)
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- Immunology (AREA)
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- Clinical Laboratory Science (AREA)
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- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)
- Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
- Automatic Analysis And Handling Materials Therefor (AREA)
Abstract
The present invention is suitable for micro-fluidic chip electrochemiluminescent immunoassay detection technique field, provides a kind of luminous micro-fluidic chip of magnetic particle, the chip includes: substrate；Sample-adding portion being disposed on the substrate, being added for sample；The storage unit being disposed on the substrate and the magnetic bead being stored in the storage unit, tagged ligand and enzyme tagged ligand；Air pump and reaction zone on the substrate is set, and the reaction zone generates with magnetic bead, tagged ligand and enzyme tagged ligand for sample and mixes and react；Cleaning solution storage unit and cleaning solution；Interconnecting piece is provided between the reaction zone and the sample-adding portion, between the reaction zone and the storage unit, between the reaction zone and the cleaning solution storage unit.The beneficial effects obtained by the present invention are as follows, by separately storing magnetic bead, tagged ligand, when use, remixes the present invention, improves the stability of magnetic bead and tagged ligand, keep the magnetic bead stored in micro-fluidic chip, tagged ligand validity period elongated, to improve the accuracy of detection.
Description
Technical field
The invention belongs to micro-fluidic chip electrochemiluminescent immunoassay detection technique field more particularly to a kind of magnetic particle shine it is micro-fluidic
Chip and reaction method.
Background technique
Currently, there are mainly two types of development trends for in-vitro diagnosis (IVD): one is automations, integrated integration, i.e., using big
Full-automatic, the highly sensitive large-scale instrument and equipment of matched central laboratory, type hospital realizes that high-precision diseases analysis is examined
Disconnected, the reagent of use is big packaging reagent, for multiple sample analysis；Another kind miniaturization, bed sideization analyzer, using list
Person-portion packs reagent, realizes the quick analyzing and diagnosing in scene.
Small hospital or community hospital's insufficient funds, sample size are few, are not appropriate for the large scale equipment of purchasing price valuableness, and
The sample for needing to analyze is few, and packs greatly after reagent is sealed off using limited time, leads to the expired waste of reagent.And point minimized
Analyzer can solve that small hospital or community hospital's large scale equipment are at high cost, reagent waste ask using single packaged reagent
Topic.
Micro-fluidic chip is also known as chip lab (Lab-on-a-chip), refers to fields such as biology, chemistry and medicine
Involved in the basic operation units such as sample preparation, reaction, separation, detection be integrated into one piece with micro-meter scale microchannel
On chip, it is automatically performed the overall process of reaction and analysis.Based on micro-fluidic chip realize analysis and detection device the advantages of be: sample
This dosage is few, and analysis speed is fast, convenient for portable instrument is made, is highly suitable for instant, field assay.
But the magnetic standard configuration body in existing single packaged micro-fluidic chip is unstable, validity period shortens, to some
When specific project is detected, test result is set to generate deviation using expired magnetic standard configuration cognition.
Summary of the invention
The embodiment of the present invention provides a kind of luminous micro-fluidic chip of magnetic particle, it is intended to solve asking for magnetic standard configuration body stability difference
Topic.
The embodiments of the present invention are implemented as follows, provides a kind of luminous micro-fluidic chip of magnetic particle, and the chip includes:
Substrate；
Sample-adding portion being disposed on the substrate, being added for sample；
The storage unit being disposed on the substrate and the magnetic bead being stored in the storage unit, tagged ligand and enzyme tagged ligand；
Air pump on the substrate is set；
The reaction zone for being disposed on the substrate, being connected to the sample-adding portion, the reaction zone are matched for sample and magnetic bead, label
Body and enzyme tagged ligand, which generate, to be mixed and reacts；
Cleaning solution storage unit and the cleaning solution being stored in the cleaning solution storage unit；
Between the reaction zone and the sample-adding portion, between the reaction zone and the storage unit, the reaction zone and institute
It states and is provided with interconnecting piece between cleaning solution storage unit.
Further, the storage unit includes:
Store the first storage chamber of the magnetic bead；
Store the second storage chamber of the tagged ligand；
Store the third storage chamber of the enzyme tagged ligand.
Further, the storage unit include store the magnetic bead, the first mixing of enzyme tagged ligand and is deposited at storage chamber
Store up the 4th storage chamber of the tagged ligand.
Further, the enzyme tagged ligand includes the ligand of enzyme or luminous agent label；
The enzyme includes: one of horseradish peroxidase and alkaline phosphatase or a variety of；The luminous agent includes: acridine
One of ester, ABEI, fluorescent dye, fluorescin and fluorescent microsphere are a variety of.
Further, the chip further include:
The detection zone being connected to the reaction zone；
Luminescent solution storage unit and the luminescent solution being stored in luminescent solution storage unit；
Between the reaction zone and the detection zone, the company of being provided between the luminescent solution storage unit and the detection zone
Socket part.
Further, the sample-adding portion, storage unit, reaction zone, detection zone, cleaning solution storage unit, luminescent solution storage unit
And micro- channel structure is used at each interconnecting piece at least one, it is micro-meter scale that micro- channel is at least one-dimensional.
The embodiment of the present invention also provides the reaction method that a kind of magnetic particle shines micro-fluidic chip, and the reaction zone includes the
One reaction subregion and the second reaction subregion, which comprises
Sample is added from sample-adding portion and sample is made to enter the first reaction subregion；
Enzyme tagged ligand is put into the first reaction subregion, keeps the sample and enzyme tagged ligand anti-first by air pump
It answers in subregion and is sufficiently mixed, reacts, obtain tagged ligand compound；
Tagged ligand compound is put into the second reaction subregion；
Tagged ligand, magnetic bead interval predetermined time are sequentially placed into the second reaction subregion, the label is made by air pump
Ligand complex and tagged ligand, magnetic bead react in subregion second to be sufficiently mixed, reacts, and resulting composite is obtained；
Resulting composite after cleaning reaction.
Further, the reaction zone is additionally provided with cleaning area, and the compound after the cleaning reaction specifically includes:
Cleaning solution is put into the second reaction subregion, the resulting composite after cleaning reaction；Or
So that the resulting composite after reaction is entered cleaning area by air pump, and cleaning solution is put into cleaning area, cleaning is anti-
Resulting composite after answering.
The embodiment of the present invention also provides a kind of reaction method of luminous micro-fluidic chip of magnetic particle, which comprises
Sample is added from sample-adding portion and sample is made to enter reaction zone；
Magnetic bead, tagged ligand and enzyme tagged ligand in storage unit is put into reaction zone；
Sample is set to be sufficiently mixed, react in reaction zone with magnetic bead, tagged ligand and enzyme tagged ligand by air pump；
Compound after cleaning reaction.
Further, the reaction zone is additionally provided with cleaning area, and the compound after the cleaning reaction specifically includes:
Cleaning solution is put into reaction zone, the compound after cleaning reaction；Or
So that the compound after reaction is entered cleaning area by air pump, and cleaning solution is put into cleaning area, after cleaning reaction
Compound.
The beneficial effects obtained by the present invention are as follows the present invention is by separately storing magnetic bead, tagged ligand, when use, mixes again
It closes, improves the stability of magnetic bead and tagged ligand, keep the magnetic bead stored in micro-fluidic chip, tagged ligand validity period elongated, from
And improve the accuracy of detection.
Detailed description of the invention
Fig. 1 is the detonation configuration figure of the luminous micro-fluidic chip one embodiment of magnetic particle provided in an embodiment of the present invention；
Fig. 2 is the detonation configuration figure of luminous another embodiment of micro-fluidic chip of magnetic particle provided in an embodiment of the present invention；
Fig. 3 is the flow chart of the reaction method for the luminous micro-fluidic chip of magnetic particle that the embodiment of the present invention eight provides；
Fig. 4 is the flow chart of the reaction method for the luminous micro-fluidic chip of magnetic particle that the embodiment of the present invention ten provides.
Specific embodiment
In order to make the objectives, technical solutions, and advantages of the present invention clearer, with reference to the accompanying drawings and embodiments, right
The present invention is further elaborated.It should be appreciated that the specific embodiments described herein are merely illustrative of the present invention, and
It is not used in the restriction present invention.
The present invention mixes sample, reacts, separating and detection is integrated on chip by using microfluidic chip technology,
And all reagent components needed for reaction are integrated on chip；It is easy to operate, when detection, sample only need to be added, close the lid,
Chip is put into miniature portable necessary instrument.
Embodiment one
As shown in Figure 1 and Figure 2, the embodiment of the present invention provides a kind of luminous micro-fluidic chip of magnetic particle, and the chip includes:
Substrate；
Sample-adding portion 21 being disposed on the substrate, being added for sample；
The storage unit being disposed on the substrate and the magnetic bead being stored in the storage unit, tagged ligand and enzyme tagged ligand；
Air pump 22 on the substrate is set；
The reaction zone 24 for being disposed on the substrate, being connected to the sample-adding portion 21, the reaction zone is for sample and magnetic bead, mark
Note ligand and enzyme tagged ligand, which generate, to be mixed and reacts；
Cleaning solution storage unit 25 and the cleaning solution being stored in the cleaning solution storage unit 25；
Between the reaction zone and the sample-adding portion 21, between the reaction zone and the storage unit, the reaction zone with
Interconnecting piece 27 is provided between the cleaning solution storage unit 25.
By separately storing magnetic bead, tagged ligand, when use, remixes the present invention, improves the steady of magnetic bead and tagged ligand
It is qualitative, keep the magnetic bead stored in micro-fluidic chip, tagged ligand validity period elongated, to improve the accuracy of detection.
Specifically, tagged ligand includes temperature-sensitive material and solution, and it is equal that magnetic bead generates liquid after mixing with tagged ligand
Phase magnetic standard configuration body.Wherein, magnetic bead includes one of Streptavidin MagneSphere, antibody modification magnetic bead and Modified antigen magnetic bead or more
Kind；Temperature-sensitive material is heat-convertible gel, includes gelatin, agar, alginate, carragheen, hydroxymethyl cellulose, Arab
One of glue, guar gum, the ashamed bean gum of thorn, pectin, starch and xanthan gum are a variety of；Solution is to contain surfactant, egg
The buffer system of white matter.After magnetic bead is mixed with temperature-sensitive material and solution, the homogeneous magnetic standard configuration body of liquid is formed, due to liquid
Good fluidity when detection, is sufficiently mixed the homogeneous magnetic standard configuration body of liquid, enzyme tagged ligand and the sample three of addition, energy
The speed of reaction is enough effectively improved, to improve the sensitivity of detection, repeatability and accuracy.
Specifically, the cleaning solution removes analyte, the luminous agent marker of non-specific adsorption for cleaning magnetic bead
And the substance of other influences testing result.Cleaning solution mainly includes buffer system, protein and surfactant, wherein buffer body
System is 6.0~10.0 including but not limited to borate, phosphate, Tris-HCl and acetate etc., the pH range of cleaning solution.Wherein
Protein is including but not limited to bovine serum albumin(BSA), casein etc..Wherein surface-active including but not limited to may include polysorbas20,
Tween 80, triton x-100, polyethylene glycol and polyvinylpyrrolidone etc..Preferably, using cleaning in the present embodiment
Liquid is the pH7.0Tris-HCl buffer comprising bovine serum albumin(BSA), tween 20 and Proclin300.
Specifically, the storage unit is seal chamber, sealing material used uses elastic material or high-isolation film, specially
Glass, plastics, rubber, aluminium foil or high-isolation film, it can also be multiple material group that wherein sealing material can form for same material
It closes.Under physical impact, storage unit can partial fracture, so that the material of storage is released.
Specifically, identical or different material and method production can be used in the storage unit and cleaning solution storage unit 25.It is preferred that
, storage unit and cleaning solution storage unit 25 is all made of plastics and elastic rubber is sealed to form.Another is preferred, and storage unit uses
Plastics and elastic rubber are sealed to form, and cleaning solution storage unit 25 is sealed to form using high-isolation film.
Air pump 22 is the air bag being built on the top plate 11 of substrate, which is connected to interconnecting piece 27, by repeatedly extruding
Or release air bag, so that the air in air bag is passed in and out interconnecting piece 27 repeatedly, to drive the liquid in top plate 11.The air pump 22 is used for
The air for absorbing or being press-connected portion 27 makes the homogeneous magnetic standard configuration body of liquid, liquid tagged ligand and sample flow to the first reaction
In subregion 241, and the air by absorbing or being press-connected portion 27 repeatedly, make the homogeneous magnetic standard configuration body of liquid and liquid tagged ligand
It reacts in subregion 241 and is sufficiently mixed first with sample.But 22 need of work of air pump is realized in sealed environment, in order to make
Chip interior sealing, is additionally provided with sealing cover (not shown), sample is added to sample-adding portion at the adding mouth in sample-adding portion 21
After in 21, then cover sealing cover.And the liquid of bottom plate 12, then it drives through capillary action, makes liquid from the second reaction subregion 242
Flow detection area 28.
Specifically, the present invention is by magnetic bead coating, dry in storage unit, and devises magnet active drive magnetic bead and (and pass
System micro-fluidic chip generally uses fluid driving or electric drive), to make magnetic bead redissolve, and real in micro-fluidic chip different zones
It is now immunoreacted, cleans, shines.Easily precipitating, poor repeatability etc. when this design not only solves magnetic bead applied to micro-fluidic chip
Problem also achieves more controllable immune response and physical cleaning, improves sensitivity and repeatability.Wherein magnet magnetism and magnetic
Pearl size significantly affects detection effect, and select magnet magnetic induction intensity of the present invention is 500-30000 Gauss, preferably 1000-
8000 Gausses；Magnetic bead is having a size of 0.1-10 μm, and preferably 0.5-3 μm.
Embodiment two
On the basis of example 1, the storage unit of the present embodiment includes:
Store the first storage chamber 231 of the magnetic bead；
Store the second storage chamber 232 of the tagged ligand；
Store the third storage chamber 233 of the enzyme tagged ligand.
In the present embodiment, the first storage chamber 231, the second storage chamber 232, third storage chamber 233 can be used identical or different
Material and method production.Preferably, the first storage chamber 231, the second storage chamber 232, third storage chamber 233 are all made of plastics and bullet
Property rubber seal forms.Another is preferred, and the first storage chamber 231 is sealed to form using plastics and elastic rubber, and second deposits
Storage chamber 232, third storage chamber 233 are sealed to form using high-isolation film.Magnetic bead, tagged ligand and enzyme tagged ligand are separately stored up
It deposits, the performance for capableing of each material of effective guarantee is independent, does not interact.
Embodiment three
On the basis of example 1, the storage unit of the present embodiment include store the magnetic bead, enzyme tagged ligand first
It mixes storage chamber and stores the 4th storage chamber of the tagged ligand.
In the present embodiment, when both the magnetic bead of selection and enzyme tagged ligand mixed stability, magnetic bead and enzyme can be marked
Both ligands are in the first mixing storage chamber, and the first mixing storage chamber is sealed to form using plastics and elastic rubber, by magnetic bead
Storage is mixed with both enzyme tagged ligands, reduces mixing step, it is simple and convenient convenient for the quick progress of subsequent reactions.
Example IV
On the basis of one to three any embodiment of embodiment, the liquid tagged ligand includes matching for luminous agent label
Body.
The luminous agent can also include: one of acridinium ester, ABEI, fluorescent dye, fluorescin and fluorescent microsphere
Or it is a variety of.
The cleaning solution storage unit 25 and the cleaning solution being stored in cleaning solution storage unit 25；
The detection zone 28 being connected to the cleaning area 29；
Luminescent solution storage unit 26 and the luminescent solution being stored in luminescent solution storage unit 26, for further cleaning magnetic bead or increasing
Strong luminous signal；
Between the cleaning area 29 and the detection zone 28, between the luminescent solution storage unit 26 and the detection zone 28
It is provided with interconnecting piece 27.
Specifically, the ligand includes: one of antigen, antibody, haptens and nucleic acid or a variety of.
Luminous agent is in conjunction with analyte or competes, and forms luminous agent tagged ligand；Magnetic particle marker in conjunction with analyte or
Competition, forms marked by magnetic bead ligand, both described ligands may be the same or different；The magnetic marker ligand, luminous agent label are matched
The ligand that body uses include nucleic acid, antigen, monoclonal antibody, polyclonal antibody and hormone receptor, the analyte include DNA,
Small molecule (drug or drugs), antigen, antibody, hormone, antibiotic, bacterium or virus and other biochemical markers.
In the present embodiment, the liquid tagged ligand can with the homogeneous magnetic mark ligand binding (such as double antibody sandwich method) of liquid or
Person and tagged ligand competition (such as competition law).Wherein the ligand of luminous agent label can be identical as the homogeneous magnetic standard configuration body of liquid,
It can be different.Preferably, in one embodiment of the invention, selecting two kinds of different antibodies as liquid tagged ligand and liquid
The homogeneous magnetic standard configuration body of state tests and analyzes object with double antibody sandwich method.
Embodiment five
On the basis of one to three any embodiment of embodiment, the enzyme tagged ligand includes the ligand of enzyme label；
The enzyme includes: one of horseradish peroxidase and alkaline phosphatase or a variety of.
Specifically, the ligand includes: one of antigen, antibody, haptens and nucleic acid or a variety of.
Enzyme or luminous agent in conjunction with analyte or compete, and form enzyme tagged ligand；Magnetic particle marker in conjunction with analyte or
Competition, forms marked by magnetic bead ligand, both described ligands may be the same or different；The magnetic marker ligand, enzyme tagged ligand make
Ligand includes nucleic acid, antigen, monoclonal antibody, polyclonal antibody and hormone receptor, and the analyte includes DNA, small point
Sub (drug or drugs), antigen, antibody, hormone, antibiotic, bacterium or virus and other biochemical markers.
In the present embodiment, the enzyme tagged ligand (such as double antibody sandwich method) or can match in conjunction with tagged ligand with label
Body competes (such as competition law).Wherein the ligand of enzyme label can be identical as tagged ligand, can also be different.Preferably, at this
In one embodiment of invention, two kinds of different antibodies is selected to detect as enzyme tagged ligand and tagged ligand with double antibody sandwich method
Analyte.In another embodiment of the invention, a kind of antigen and a kind of antibody are selected, respectively as enzyme tagged ligand and label
Ligand detects sample with competition law.
Embodiment six
In an alternative embodiment of the invention, the chip further include:
The detection zone 28 being connected to the reaction zone；
Luminescent solution storage unit 26 and the luminescent solution being stored in luminescent solution storage unit 26；
It is all provided between the reaction zone and the detection zone 28, between the luminescent solution storage unit 26 and the detection zone 28
It is equipped with interconnecting piece 27.
Specifically, the detection zone 28 is connected to the cleaning area 29.
Cleaning solution storage unit 25 is set as being connected to cleaning area 29 or being connected to reaction zone 24.
In the present embodiment, the enzyme uses one of horseradish peroxidase and alkaline phosphatase or a variety of；It is described to shine
Agent uses one of acridinium ester, ABEI, fluorescent dye, fluorescin and fluorescent microsphere or a variety of；Carry out chemiluminescence needs
Luminescent solution, and light emitting source is then needed using fluorescence radiation.
Detection zone 28 is set, convenient for resulting composite being moved to detection zone 28 and is seen after cleaning solution being cleaned
It examines and detects, be easy to use, it is simple and convenient.
Specifically, the substrate is additionally provided with the waste liquid storage unit 30 being connected to the cleaning area.Convenient for collection cleaning and instead
Waste liquid after answering can reduce interference of the waste liquid to detection, effectively improve the precision of detection.
Embodiment seven
In an alternate embodiment of the present invention where, the sample-adding portion 21, storage unit, reaction zone 24, detection zone 28, cleaning
Micro- channel structure is used at liquid storage unit 25, luminescent solution storage unit 26 and each interconnecting piece 27 at least one, micro- channel is extremely
One-dimensional less is micro-meter scale.
Micro-fluidic chip of the invention will test all reagent components needed for process, and (enzyme standard configuration body, magnetic bead, label are matched
Body, cleaning solution, luminescent solution etc.) it integrates, be built into micro-fluidic chip, and by ingenious micro- channel design, in necessary instrument
Operation under, realize the operating in a key (only need to can be achieved with detecting by start button, be not necessarily to complex operations) of micro-fluidic chip, it is real
Existing whole blood separation, immune response, cleaning separation, chemiluminescence detection, are designed so as to avoid structure in existing micro-fluidic chip
Simply, the deficiencies of complicated for operation when detecting and defect.Serum or blood plasma detection can only be carried out by also overcoming traditional chemical light-emitting appearance,
And the shortcomings that whole blood sample cannot being detected.
Embodiment eight
As shown in Figure 1, in an alternative embodiment, the substrate includes:
Top plate 11；
The top adhesive tape 13 at 11 top of top plate is set；
The bottom tape 14 of 11 bottom of top plate is set；
The sample-adding portion 21, storage unit, reaction zone and each interconnecting piece 27 are arranged on the top plate 11.
In the present embodiment, the moulding material of the substrate is polymer, including but not limited to polystyrene, polyvinyl chloride,
Polypropylene, epoxy resin etc..
As shown in Fig. 2, in another alternative embodiment, the substrate includes top plate 11 and bottom plate 12；
The storage unit, reaction zone and each interconnecting piece 27 are provided on the top plate 11 and bottom plate 12.
The sample-adding portion 21 is arranged on the top plate 11.
In the present embodiment, the moulding material of the substrate is polymer, including but not limited to polystyrene, polyvinyl chloride,
Polypropylene, epoxy resin etc..
The embodiment of the present invention uses microfluidic chip technology, and sample is mixed, is reacted, is separated and detection is integrated in chip
On, and all reagent components needed for reaction are integrated on chip.It is easy to operate, when detection, sample only need to be added, close the lid
Son is put into chip in miniature portable necessary instrument.
Specifically, the top adhesive tape 13 is equipped with the adding mouth 134 being used cooperatively with sample-adding portion 21, the adding mouth 134
Place is equipped with sealing cover 133.The top plate 11 is equipped with the second window for being placed on cleaning solution storage unit 25,26 top of luminescent solution storage unit
Mouth 132, and the first window 131 of the first storage chamber 231 of setting, 232 top of the second storage chamber, are arranged first window 131, second
Window 132 squeezes each deposit convenient for making the pressure ram of analyzer put in chip interior from first window 131, the second window 132
Storage portion, so that the liquid substance in each storage unit flows out；And sealing cover 133 is set, convenient for sealing sample-adding portion 21, to make
It obtains chip interior and keeps sealing, work convenient for air pump.
Embodiment nine
As shown in figure 3, the embodiment of the present invention also provides a kind of reaction method of luminous micro-fluidic chip of magnetic particle, it is described anti-
Answering area includes the first reaction subregion 241 and the second reaction subregion 242, which comprises
Sample is added from sample-adding portion 21 and sample is made to enter the first reaction subregion 241；
Enzyme tagged ligand is put into the first reaction subregion 241, so that the sample and enzyme tagged ligand is existed by air pump 22
It is sufficiently mixed, reacts in first reaction subregion 241, obtain tagged ligand compound；
Tagged ligand compound is put into the second reaction subregion 242；
Tagged ligand, magnetic bead interval predetermined time are sequentially placed into the second reaction subregion 242, made by air pump 22 described
Tagged ligand compound and tagged ligand, magnetic bead react in subregion 242 second to be sufficiently mixed, reacts, and resulting composite is obtained；
Resulting composite after cleaning reaction.
In the present invention, the first reaction subregion 241 can be one for reaction type chamber, be also possible to shared interconnecting piece 27
Some.
In the present embodiment, magnetic bead, tagged ligand can combine (liquid or solid-state) by particular combination power；The enzyme label is matched
Body can in conjunction with tagged ligand (such as double antibody sandwich method) or with tagged ligand compete (such as competition law).
By separately storing magnetic bead, tagged ligand, when use, remixes the present invention, improves the steady of magnetic bead and tagged ligand
It is qualitative, keep the magnetic bead stored in micro-fluidic chip, tagged ligand validity period elongated, to improve the accuracy of detection.
Embodiment ten
On the basis of embodiment nine, the reaction zone is additionally provided with cleaning area 29, the compound tool after the cleaning reaction
Body includes:
Cleaning solution is put into the second reaction subregion 242, the resulting composite after cleaning reaction；Or
So that the resulting composite after reaction is entered cleaning area by air pump 22, and cleaning solution is put into cleaning area, cleans
Resulting composite after reaction.
In the present embodiment, magnetic bead is cleaned by cleaning solution, remove the analyte of non-specific adsorption, luminous agent marker and
The substance of other influences testing result；The precision of detection can be further increased.
Embodiment 11
As shown in figure 4, the embodiment of the present invention also provides a kind of reaction method of luminous micro-fluidic chip of magnetic particle, the side
Method includes:
Sample is added from sample-adding portion 21 and sample is made to enter reaction zone；
Magnetic bead, tagged ligand and enzyme tagged ligand in storage unit is put into reaction zone；
Sample is set to be sufficiently mixed, react in reaction zone with magnetic bead, tagged ligand and enzyme tagged ligand by air pump 22；
Compound after cleaning reaction.
In the present embodiment, magnetic bead, tagged ligand can combine (liquid or solid-state) by particular combination power；The enzyme label is matched
Body can in conjunction with tagged ligand (such as double antibody sandwich method) or with tagged ligand compete (such as competition law).
By separately storing magnetic bead, tagged ligand, when use, remixes the present invention, improves the steady of magnetic bead and tagged ligand
It is qualitative, keep the magnetic bead stored in micro-fluidic chip, tagged ligand validity period elongated, to improve the accuracy of detection.
Embodiment 12
On the basis of embodiment 11, the reaction zone is additionally provided with cleaning area, the compound tool after the cleaning reaction
Body includes:
Cleaning solution is put into reaction zone, the compound after cleaning reaction；Or
So that the compound after reaction is entered cleaning area by air pump 22, and cleaning solution is put into cleaning area, cleaning reaction
Compound afterwards.
In the present embodiment, magnetic bead is cleaned by cleaning solution, remove the analyte of non-specific adsorption, luminous agent marker and
The substance of other influences testing result；The precision of detection can be further increased.
By separately storing magnetic bead, tagged ligand, when use, remixes the present invention, improves the steady of magnetic bead and tagged ligand
It is qualitative, keep the magnetic bead stored in micro-fluidic chip, tagged ligand validity period elongated, to improve the accuracy of detection.
Specifically, magnetic bead, tagged ligand and enzyme tagged ligand are separately stored, it is capable of the performance of each material of effective guarantee
It is independent, it does not interact.The mixing storage chamber is sealed to form using plastics and elastic rubber, and three is mixed and is stored, is convenient for
The quick progress of subsequent reactions, it is simple and convenient.The enzyme tagged ligand can in conjunction with tagged ligand (such as double antibody sandwich method) or
Person and tagged ligand competition (such as competition law).The ligand that wherein ligand of luminous agent label is marked with enzyme can be with tagged ligand phase
Together, it can also be different.Preferably, in one embodiment of the invention, selecting two kinds of different antibodies as enzyme tagged ligand
Object is tested and analyzed with double antibody sandwich method with tagged ligand.In another embodiment of the invention, a kind of antigen and one kind are selected
Antibody detects sample respectively as enzyme tagged ligand and tagged ligand with competition law.Detection zone 28 is set, convenient in cleaning solution
After being cleaned, resulting composite is moved to detection zone 28 and is observed and is detected, is easy to use, it is simple and convenient.The present invention
Micro-fluidic chip will test all reagent component (enzyme standard configuration body, magnetic bead, tagged ligand, cleaning solutions, luminescent solution needed for process
Deng) integrate, be built into micro-fluidic chip, and by ingenious micro- channel design, under the operation of necessary instrument, realize miniflow
The operating in a key (only need to can be achieved with detecting by start button, be not necessarily to complex operations) for controlling chip, realizes that whole blood separates, is immune anti-
It answers, clean separation, chemiluminescence detection, operation is multiple when, detection simple so as to avoid structure design in existing micro-fluidic chip
The deficiencies of miscellaneous and defect.Serum or blood plasma detection can only be carried out by also overcoming traditional chemical light-emitting appearance, and cannot be to whole blood sample
The shortcomings that being detected.
The present invention is not simple superposition magnetic microparticle chemiluminescence technology and microfluidic chip technology, but passes through hydraulic seal
Design, channel design, integrate the required all chemical constituents of detection, are built into micro-fluidic chip, and with magnet active drive,
Realize one-touch magnetic microparticle chemiluminescence immune detection, thus in portable necessary instrument realize whole blood in analyte it is fast
Speed, highly sensitive, accurate quantitative analysis detection.
The foregoing is merely illustrative of the preferred embodiments of the present invention, is not intended to limit the invention, all in essence of the invention
Made any modifications, equivalent replacements, and improvements etc., should all be included in the protection scope of the present invention within mind and principle.
Claims (10)
- The micro-fluidic chip 1. a kind of magnetic particle shines, which is characterized in that the chip includes:Substrate；Sample-adding portion being disposed on the substrate, being added for sample；The storage unit being disposed on the substrate and the magnetic bead being stored in the storage unit, tagged ligand and enzyme tagged ligand；Air pump on the substrate is set；The reaction zone for being disposed on the substrate, being connected to the sample-adding portion, the reaction zone for sample and magnetic bead, tagged ligand and Enzyme tagged ligand, which generates, to be mixed and reacts；Cleaning solution storage unit and the cleaning solution being stored in the cleaning solution storage unit；Between the reaction zone and the sample-adding portion, between the reaction zone and the storage unit, the reaction zone and described clear Interconnecting piece is provided between washing lotion storage unit.
- The micro-fluidic chip 2. magnetic particle as described in claim 1 shines, which is characterized in that the storage unit includes:Store the first storage chamber of the magnetic bead；Store the second storage chamber of the tagged ligand；Store the third storage chamber of the enzyme tagged ligand.
- The micro-fluidic chip 3. magnetic particle as described in claim 1 shines, which is characterized in that the storage unit includes described in storage 4th storage chamber of magnetic bead, the first mixing storage chamber of enzyme tagged ligand and the storage tagged ligand.
- The micro-fluidic chip 4. magnetic particle as described in any one of claims 1-3 shines, which is characterized in that the enzyme tagged ligand The ligand marked including enzyme or luminous agent；The enzyme includes: one of horseradish peroxidase and alkaline phosphatase or a variety of；The luminous agent include: acridinium ester, One of ABEI, fluorescent dye, fluorescin and fluorescent microsphere are a variety of.
- The micro-fluidic chip 5. magnetic particle as claimed in claim 4 shines, which is characterized in that the chip further include:The detection zone being connected to the reaction zone；Luminescent solution storage unit and the luminescent solution being stored in luminescent solution storage unit；Connection is provided between the reaction zone and the detection zone, between the luminescent solution storage unit and the detection zone Portion.
- The micro-fluidic chip 6. magnetic particle as claimed in claim 5 shines, which is characterized in that the sample-adding portion, storage unit, reaction Micro- channel structure is used at area, detection zone, cleaning solution storage unit, luminescent solution storage unit and each interconnecting piece at least one, it is described It is micro-meter scale that micro- channel is at least one-dimensional.
- 7. a kind of reaction method using the luminous micro-fluidic chip of the described in any item magnetic particles of claim 1~6, the reaction Area includes the first reaction subregion and the second reaction subregion, which is characterized in that the described method includes:Sample is added from sample-adding portion and sample is made to enter the first reaction subregion；Enzyme tagged ligand is put into the first reaction subregion, reacts the sample and enzyme tagged ligand point first by air pump It is sufficiently mixed, reacts in area, obtain tagged ligand compound；Tagged ligand compound is put into the second reaction subregion；Tagged ligand, magnetic bead interval predetermined time are sequentially placed into the second reaction subregion, the tagged ligand is made by air pump Compound and tagged ligand, magnetic bead react in subregion second to be sufficiently mixed, reacts, and resulting composite is obtained；Resulting composite after cleaning reaction.
- 8. reaction method as claimed in claim 7, which is characterized in that the reaction zone is additionally provided with cleaning area, and the cleaning is anti- Compound after answering specifically includes:Cleaning solution is put into the second reaction subregion, the resulting composite after cleaning reaction；OrSo that the resulting composite after reaction is entered cleaning area by air pump, and cleaning solution is put into cleaning area, after cleaning reaction Resulting composite.
- 9. a kind of reaction method using the luminous micro-fluidic chip of the described in any item magnetic particles of claim 1~6, feature exist In, which comprisesSample is added from sample-adding portion and sample is made to enter reaction zone；Magnetic bead, tagged ligand and enzyme tagged ligand in storage unit is put into reaction zone；Sample is set to be sufficiently mixed, react in reaction zone with magnetic bead, tagged ligand and enzyme tagged ligand by air pump；Compound after cleaning reaction.
- 10. reaction method as claimed in claim 9, which is characterized in that the reaction zone is additionally provided with cleaning area, and the cleaning is anti- Compound after answering specifically includes:Cleaning solution is put into reaction zone, the compound after cleaning reaction；OrSo that the compound after reaction is entered cleaning area by air pump, and cleaning solution is put into cleaning area, answering after cleaning reaction Close object.
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