CN106996973A - A kind of sample testing apparatus - Google Patents
A kind of sample testing apparatus Download PDFInfo
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- CN106996973A CN106996973A CN201710274311.0A CN201710274311A CN106996973A CN 106996973 A CN106996973 A CN 106996973A CN 201710274311 A CN201710274311 A CN 201710274311A CN 106996973 A CN106996973 A CN 106996973A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/558—Immunoassay; Biospecific binding assay; Materials therefor using diffusion or migration of antigen or antibody
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Abstract
The present invention relates to a kind of sample testing apparatus, including test strips, test strips include sample pad, label pad and detecting pad, label pad is located at the downstream of sample pad, and detecting pad is located at the downstream of label pad, and the sample testing apparatus also includes collecting chamber, capture pad and body, capture pad is located in collecting chamber, test strips are located within the body, the test strips liquid communication in capture pad and body in collecting chamber, and capture pad is positioned at the upstream of test strips fluid course.The device of the detection sample of the present invention is because the antibody on capture pad and capture pad is divided into two parts and separates, add the time of the combination of analyte and space in antibody and sample, so that with reference to more abundant, especially for special sample, the possibility of the false positive of exclusion, improves the accuracy of detection.
Description
The application is point proposed in the application in original application day on July 1st, 2014 in original applying number 201410309729.7
Case application.
Technical field
The present invention relates to the device of detection sample, the particularly test strips for detecting sample.
Background technology
Detect that this technology in sample with the presence or absence of analyte is used extensively using immune association reaction principle
In every field.The analyte of various biological specimens (saliva, blood, urine, serum, sweat etc.) can be detected with it
Matter comes monitoring of diseases and the health status (early pregnancy, tumour, infectious disease, drugs etc.) of the mankind.The basic original of this detection technique
Reason is built upon the performance between immune molecule with specific bond, such as antibody and antigen, haptens/antibody, biotin with
Antibiotin etc..In addition, many such detections can be completed on solid dielectric, such as conventional lateral flow reagent
In bar, glass or plastic microtiter plates, device for immunochromatography etc..Generally, can be in conjunction with one on immune specific binding molecule
A little solid particles or chemical substance, so can come qualitative by naked eyes or other instruments equipment, quantitatively or semi-quantitatively must
Go out testing result.This solid particle can be colored colloidal solid (latex or gold grain), and this chemical substance can be with
It is the material with chromophoric group, these materials can send specific wavelength to show detection knot under the conditions of other are suitable
Really.
It can be found in the prior art using the detection reagent bar or device of these principles, such as the following patent
The reagent strip of description or the device containing reagent strip：US 4857453；US 5073484；US 5119831；US 5185127；US
5275785；US 5416000；US 5504013；US 5602040；US 5622871；US 5654162；US 5656503；US
5686315；US 5766961；US 5770460；US 5916815；US 5976895；US 6248598；US 6140136；US
6187269；US 6187598；US 6228660；US 6235241；US 6306642；US 6352862；US 6372515；US
6379620；With US 6403383.
Immune detection generally includes two kinds of principles, sandwich and competition law, wherein being detected with competing method in sample
Haptens small-molecule substance it is most commonly seen.The method and reagent and detection means detected using competition law is in United States Patent (USP)
US4235601；It is described later in detail in US4442204, US5208535, US5229073.These devices are all descriptions in detection
When not having color change or the no colored line to occur on region or in result reading area, testing result is judged to the positive,
Represent that detection sample there may be analyte, on the contrary, when occur in detection zone or result reading area color change or
When person has the colored line to occur, testing result is judged to feminine gender, represents that analyte may be not present in detection sample.
In some detection means, sample process region is separated with test strips, such as Chinese patent
A kind of detection means described in CN200610052628.1, including second reagent areas.Second reagent areas and examination
Paper slip is separated, and is shifted to an earlier date and sample contact, the analyte in sample is sufficiently combined with its antibody；Then, it is processed
The sample crossed flows to test strips and detected again.
In actual operation, there is problems with this detection means：For it is sticky or more than the low foam of water content
Most early stage can not be captured the antibody in pad (the second reagent areas) and be eluted completely by saliva sample, i.e. sample can not be with
Antibody on capture pad is sufficiently combined, and causes the antigen binding in antibody and label pad without sufficient amount, therefore nothing
Method develops the color, so as to cause false positive.
The content of the invention
In order to solve these problems, the present invention provides an improved detection means and the side using the improved device
Method, the analyte and the molecule of the specific bond on capture pad in sample can be made by improved detection means and method
(antibody) is more fully combined, so that the problem of effectively overcoming false positive, improves the accuracy rate of Product checking.
On the one hand, it is provided by the present invention it is a kind of detect sample device, including, test strips, test strips include sample pad,
Label pad and detecting pad, label pad are located at the downstream of label pad, and detecting pad is located at the downstream of label pad；Characterized in that, the inspection
The device of test sample sheet also includes capture pad, and the capture pad is located at the upstream of test strips and circulated with test strips liquid phase；The capture
Pad can be connected or be not connected to test strips.
One preferred embodiment in, the capture pad includes the first capture pad (capture pad) and the second capture pad (is buffered
Pad).
In the mode being more highly preferred to, the first capture pad (capture pad) is located at second capture pad (cushion pad) upstream, and the two is not
It is connected, but liquid communication.
In one specific embodiment, the second capture pad (cushion pad) is located at the upstream of test strips, the two and liquid phase
Connection.Now, the second capture pad (cushion pad) can be connected with test strips, can also be not connected with.
Capture pad includes two, and is not connected with, and lengthens the time that sample flows through, i.e. total stop on capture pad
Time lengthens.When including the reagent reacted with analyte in sample on capture pad, analyte and examination can be made
Agent has more fully time and space to be combined.
In some embodiments, the second capture pad (cushion pad) is located at the upstream of the sample pad of test strips, with sample pad phase
Connection.
In one specific embodiment, the second capture pad (cushion pad) is the prolongation of sample pad.
In other embodiments, capture mat material matter is polyester fiber film or glass fibre membrane.
In preferred embodiment, the first capture pad (capture pad) material is polyester fiber film；Second capture pad (buffering
Pad) material be glass fibre membrane.
In some embodiments, point comprising a kind of specific bond analyte that can be flowed with liquid on capture pad
Son.
In other embodiments, capture pad also include with the incoherent specific binding molecule of analyte to one kind
Molecule.
The first capture pad (capture pad) and the second capture pad on (cushion pad) for the reagent that is combined with analyte
(molecule of specific bond analyte and with the incoherent specific binding molecule of analyte to a kind of molecule) have
After certain proportioning, can more fully it be reacted with the analyte in sample.Some preferred embodiment in, second
Capture pad (cushion pad) on specific bond analyte molecule and with the incoherent specific binding molecule of analyte
To the content of molecule a kind of the 15%-50% of (cushion pad) total amount is padded for the first capture pad (capture pad) and the second capture.
One more specifically in embodiment, the molecule of the specific bond analyte on the second capture pad (cushion pad)
And with the incoherent specific binding molecule of analyte to a kind of molecule content for the first capture pad (capture pad) and
The 20%-35% of second capture pad (cushion pad) total amount.
Include in some embodiments, on detecting pad a kind of fixed, uncorrelated to the upper analyte of capture pad
Specific binding molecule to another molecule.
In other embodiments, with the specific bond on the second capture pad (cushion pad) on the first capture pad (capture pad)
The molecule of analyte includes the first antibody of analyte；Label pad includes the secondary antibody of analyte.
It is preferred that, it is described with the incoherent specific binding molecule of analyte to selected from following molecule to one of：
Biotin/avidin (biotin/avidin)；Biotin/Streptavidin (biotin/streptavidin)；If rhodamine/
Red bright antibody (rhodamine/anti-rhodamine)；Mouse IgG/ mouse IgG antibody (Mouse IgG/anti-mouse
IgG)。
On capture pad reagent (molecule of specific bond analyte and with the incoherent spy of analyte
Different binding molecule to a kind of molecule) be processed and be fixed on capture pad, when liquid sample is added on capture pad
And when flowing through capture pad, can be flowed by reagent of the fixing process on capture pad with after analyte combination with liquid.
Wherein, reagent be processed at capture pad upper type it is various.In some specific embodiments, the molecule of specific bond analyte
And with the incoherent specific binding molecule of analyte to a kind of molecule be processed at whole first capture pad (capture
Pad) and second capture pad (cushion pad) on.In this mode, analyte can be made sufficiently to be reacted with reagent.
Other preferred embodiment in, the molecule of specific bond analyte and incoherent with analyte
Specific binding molecule to a kind of molecule be processed on whole first capture pad (capture is padded)；Specific bond analyte
Molecule and with the incoherent specific binding molecule of analyte to a kind of molecule be processed in linearly aligned mode
On one line of the second capture pad (cushion pad).Linearly aligned reagent arrangement on second capture pad (cushion pad) is more concentrated,
All samples is flowed through linear place and carry out sufficient liquid contact, make with reference to more abundant, so that also more effective
The mode false positive of product.
On the other hand, a kind of sample testing apparatus, including test strips, test strips include sample pad, label pad and detecting pad,
Label pad is located at the downstream of sample pad, and detecting pad is located at the downstream of label pad, and the sample testing apparatus also includes collecting chamber, capture
Pad and body, capture pad are located in collecting chamber, and test strips are located within the body, the test strips in capture pad and body in collecting chamber
Liquid communication, and capture pad is positioned at the upstream of test strips fluid course.
In some embodiments, sample testing apparatus also includes cushion pad, and cushion pad is located at the upper of the sample pad of test strips
Trip, and be connected with sample pad.
Some preferred embodiment in, the body also includes sample ingress area, and collecting chamber is connected with ingress area
Connect；Have on ingress area and import through hole, the importing through hole makes ingress area and test strips liquid communication in body.
In the embodiment being more highly preferred to, collecting chamber includes the first collecting chamber and the second collecting chamber, and the first collecting chamber is the
In two collecting chambers；First collecting chamber, which has, makes the first liquid through-hole of the first collecting chamber and the second collecting chamber liquid communication；The
There is second liquid through hole on two collecting chambers.
Some preferred embodiment in, collecting chamber can be rotated relative to ingress area；Before rotation, the second collecting chamber
Second liquid through hole is imported into the baseplane sealing in region, collecting chamber bottom is imported into region bottom surface and is sealed；After rotation, the
The second liquid through hole of two collecting chambers is connected with the importing through hole on ingress area, collecting chamber is connected with ingress area liquid
It is logical.
In other embodiments, capture pad is loaded with reagent, and position of the capture pad in collecting chamber causes liquid by the
One collecting chamber flow into the second collecting chamber during, or liquid be located at first or the second collecting chamber when, can with capture pad on
Reagent be in contact.
In some preferred embodiments, capture pad is connected with the first collecting chamber side wall or bottom wall, or capture is padded positioned at the
In two collecting chambers, or the first collecting chamber is run through in capture pad one end, its one end is located at the first collecting chamber, the other end is located at second
Collecting chamber；Or capture pad is located in the insulating space that the first collecting chamber bottom is formed with the second collecting chamber bottom.
On the other hand, the present invention also provides a kind of method for detecting and whether there is analyte in liquid sample, including：
A kind of device for detecting sample is provided, the detection means includes reagent strip and capture is padded, described reagent strip test strips include sample
This pad, label pad and detecting pad, label pad are located at the downstream of sample pad, and detecting pad is located at the downstream of label pad；Capture pad includes
First capture pad (capture pad) and the second capture pad (cushion pad)；First capture pad (capture pad) is located at the second capture pad (buffering
Pad) upstream, the two is not connected with but liquid phase circulates；Second capture pad (cushion pad) is located at the upstream of the sample pad of test strips,
The two cocurrent body phase connection that is connected；It is characterized in that：Adding liquid sample in (capture pad) is padded to the first capture, the liquid is allowed
Sample is first with the reagent reacting on the first capture pad (capture is padded) after -60 minutes 30 seconds；Then liquid sample is allowed to flow to successively
Two capture pads (cushion pad) and sample pad, label pad, detecting pad and the reagent reacting completion inspection padded with each in test strips
Survey；Read testing result.
In some embodiments, the reagent on capture pad includes the molecule and and analyte of specific bond analyte
The incoherent specific binding molecule of matter to a kind of molecule；The molecule is connected with the molecule of described specific bond analyte
Or combine and can be flowed with liquid.
Some preferred embodiment in, second capture pad (cushion pad) on specific bond analyte molecule and
With the incoherent specific binding molecule of analyte to the content of molecule a kind of be the first capture pad (capture pad) and second
The 15%-50% of capture pad (cushion pad) total amount.
Other preferred embodiment in, padded to the first capture after (capture pad) adding liquid sample, allow liquid sample
With the reagent reacting -30 minutes 30 seconds on the first capture pad (capture pad).
The present invention also provides a kind of kit for detecting sample, including：Detect the device of sample and collect the device of sample,
Wherein, the device of detection sample includes reagent strip, and described test strips include sample pad, label pad and detecting pad, label pad position
In the downstream of sample pad, detecting pad is located at the downstream of label pad；Wherein, collecting the device of sample includes that liquid sample can be absorbed
Collection head；Also include capture pad in the device of described detection sample, capture pad is positioned at the upstream of test strips and and test strips
Liquid phase circulates；The capture pad can be connected or be not connected to test strips.
It is preferred that, sample testing apparatus also includes the second capture pad (cushion pad)；First capture pad (capture pad) is located at the
Two capture pad (cushion pad) upstreams, the two is not connected with but liquid phase circulates；Second capture pad (cushion pad) is located at test strips
The upstream of sample pad, the two cocurrent body phase connection that is connected.
Other preferred embodiment in, include on the first capture pad (capture pad) and the second capture pad (cushion pad) special
The molecule of different combination analyte and with the incoherent specific binding molecule of analyte to a kind of molecule.
In some specific embodiments, second capture pad (cushion pad) on specific bond analyte molecule and
With the incoherent specific binding molecule of analyte to the content of molecule a kind of be the first capture pad (capture pad) and second
The 15%-50% of capture pad (cushion pad) total amount.
Beneficial effect
The device of the detection sample of the present invention is added because the antibody on capture pad and capture pad is divided into two parts and separates
The time of the combination of analyte and space in antibody and sample, so that with reference to more abundant, especially for special sample
This, the possibility of the false positive of exclusion improves the accuracy of detection.
Brief description of the drawings
Fig. 1 is the decomposition texture schematic diagram of common test strip；
Fig. 2 is the top view of common test strip；
Fig. 3 is the structural representation of the device of the detection sample before the improvement of the present invention；
Fig. 4 is the structural representation of the device of the detection sample after the improvement of the present invention；
Fig. 5 detects result schematic diagram after preceding and detection for the detection means of one concrete scheme of the present invention；
Fig. 6 is that the detection means of another concrete scheme of the invention detects result schematic diagram after preceding and detection；
Fig. 7 is that the detection means of another concrete scheme of the invention detects result schematic diagram after preceding and detection；
Fig. 8 includes the decomposing schematic representation of the detection box of the device of detection sample after improving for the present invention；
Fig. 9 is the diagrammatic cross-section for the part that Fig. 8 detects box.
Description of reference numerals：
100,300 reagent strips；101 sample pads；102 label pads；103 detecting pads；132 testing result lines；133 testing results
Control line；104 adsorptive pads；105 support chips；210 capture pads；310 first capture pads (capture pad)；320 second capture pad (bufferings
Pad)；400 detection means；500 collection devices；501 collect handle；502 collect head；4011 first collecting chambers；4012 second collect
Chamber；401 collecting chambers；First liquid through-hole of the collecting chamber of 4011-1,4011-2 first；403,403-1,403-2 sealing ring；406
Sample ingress area；406-1 samples import through hole；3101 paste-sides；3102 reagent ends；404 bodies；4041 read window；Really
Recognize chamber 407；4071 bases；4073 export through holes；4072 plugs；4043 upper plates；4045 lower plates.
Embodiment
Structure of the present invention or these used technical terms are described further below.
Test strips 100
General reagent strip, as depicted in figs. 1 and 2, reagent strip 100 include sample application zone and marked region and inspection
Region is surveyed, detection zone includes a kind of specific binding molecule.The reagent strip more optimized also includes the suction positioned at detection zone downstream
Aqua region, sample can flow along the signified direction of arrow on reagent strip.Include completing reaction on sample application zone
Necessary material, such as some buffer reagents, pretreatment sample reagent etc.；Include fluorescence labeling material in marked region,
Colloid gold label particle, or to colour gel marking particle, can also be water-soluble marker's material, these mark substances can be with
Antibody, antigen, haptens or analyte similar substance etc. are connected, fixed special knot is included in detection zone
Molecule is closed, can occur color change in detection zone by specific binding molecule to represent in sample with the presence or absence of analyzed
Material.It can also include testing result control area 133 in the downstream of detection zone.Reagent strip can be made up of following part,
Sample pad 101 and label pad 102, detecting pad 103, adsorptive pads 104, these materials, which are assembled on support chip 105, constitutes whole
Reagent strip 100 (such as Fig. 1)；Include detection zone 132 wherein on detecting pad 103, preferential scheme is under detection zone
Trip also includes testing result control area 133.Liquid sample can along the signified direction of arrow from sample pad 101 sequentially through
Label pad 102, detecting pad 103 finally reach adsorptive pads 104.The all parts for constituting reagent strip can be by absorbent material structure
Into generally, detecting pad 103 can be constituted for one of nitrocellulose filter, cellulose acetate film or nylon membrane.These constitute examination
The part and material of agent bar and to handle some conventional chemical reagent and processing method on these parts be all prior art
It is disclosed.
Capture pad 210
Include the molecule of specific bond analyte that can be flowed with liquid, 210, the capture pad on capture pad 210
Separated in the upstream of reagent strip 100 and with reagent strip 100.A scheme as shown in Figure 3, capture pad 210 is located at reagent strip 100
Sample pad 101 upstream, liquid sample firstly flow through capture pad 210 then reach reagent strip 100 on sample pad on 101 complete
Whole detection.
Detection means 400
Such as Fig. 8, shown in 9, detection means 400 includes body 404 and collection well 401, wherein reagent strip 100 or 300
In body 404, capture pad is located in collection well 401；Capture pad and reagent strip are in liquid communication state.Here institute
" the liquid communication state " said refers to that liquid can be flowed on reagent strip from collecting chamber 401, and this flowing can be due to liquid sheet
The Action of Gravity Field of body and flow, can also be by being provided with through hole between collecting chamber 401 and reagent strip 300 and flowing, while receiving
Some flow guiding structures can also be installed between collection chamber and reagent strip, so liquid can be allowed more smoothly to be flowed to from collecting chamber
On reagent strip.More specifically, include reading window 4041 and sample ingress area 406, sample ingress area on body 404
406, which include sample, imports through hole 406-1 etc., the correspondence of reading window 4041 in the detection zone and body 404 on reagent strip, examination
It is corresponding that the sample application zone (sample pad) of agent bar imports through hole with sample.The sample that through hole 406-1 is flowed into is imported by sample
The sample application zone on reagent strip can be flowed to and then detection zone is reached and complete whole detection reaction.Collecting chamber 401 includes
First collecting chamber 4011 and the second collecting chamber 4012, the one end open of the first collecting chamber 4011 be used for receive need detect sample or
Person receives the collection device with sample, and several the first liquid through-hole 4011- that liquid can be allowed to pass through are provided with an opposite end
Isosorbide-5-Nitrae 011-2 etc. (Fig. 9)；There is a vertical bar beam on the first liquid through-hole 4011-1 correspondence position, the beam one end can be fixed on
On the side wall of first collecting chamber 4011, the other end extends downward and through the first liquid through-hole 4011-1, and capture pad is located at vertical bar
Liang Shang.Capture pad one end can be fixed on the surface of vertical beam, and the other end can reach the second collecting chamber through liquid through-hole
4012 bottom, include on one end of capture pad the molecule (Y1) of specific bond analyte that can be flowed with liquid and with
The incoherent specific binding molecule of analyte plants molecule (M1) to one of (M1/M2)；The material of the capture pad can be one
A little water-absorbing material compositions, some protein substances can be handled above, such as glass fibre, filter paper, acetate fiber etc. are caught
The surface of vertical beam can be bonded at by organic nothing or machine glue by obtaining one end of pad.The capture pad can also be adhesive in the first liquid through-hole
In 4011-2.Capture pad can also other forms be present in collecting chamber 401, for example capture pad can be placed directly in the first collection
The bottom of chamber 4011, can also be placed in the insulating space formed between the first collecting chamber bottom and the second collecting chamber bottom, only
Capture pad to be placed in collecting chamber 401, can allow sample fully and its contact just can be so that in collecting chamber 401, capture is padded
210 can be fully immersed in sample, partly can also be immersed in sample.It can also include some other auxiliary on capture pad
The buffer reagent of reagent, such as cushioning effect, some protein moleculars etc..Those of ordinary skill in the art combine the present invention all
The other forms or mode being readily apparent that can serve as one embodiment of the present of invention.First collecting chamber 4011 and second is received
Collection chamber 4012 is assembled into collecting chamber 401 (Fig. 8).Second collecting chamber 4012 includes one end open, and the other end is provided with several second liquid
Body through hole, corresponding sealing ring 403,403-1,403-2 etc. are separately installed with these liquid through-holes.The similar and present apparatus
Other specific structure descriptions have specific description in U.S. Patent application 2005/0180882A1.
In specific embodiment, it can also include and the incoherent specific bond of analyte on capture pad
Molecule to a kind of molecule M1, the molecule is connected or combines with the molecule Y1 of specific bond analyte.Capturing pad can position
In the upstream of sample pad on reagent strip.More specifically, it is exactly that capture is padded positioned at the upstream of sample pad 201, sample pad is located at mark
Remember the upstream of pad 202, label pad is located at the upstream of detecting pad；Specific bond is fixed with detection zone on detecting pad to be divided
Another molecule Y2 of material is analysed, mark substance L is included in label pad and is divided with the incoherent specific bond of analyte
Son to another molecule M2 (Fig. 5 A).When being reacted, if containing analyte A in sample, it is analyzed
Specific binding molecule Y1 in substance A and capture pad is combined to form compound substance A-Y1-M1；When the compound substance passes through sample
This pad is reached in label pad, just forms new compound substance A-Y1-M1-M2-L；When new compound substance is flowed through on detecting pad
When, the new compound substance of the specific binding molecule Y2 specific bonds that are fixed in detection zone, so as to go out in detection zone
Existing color represents positive findings (Fig. 5 B).
In another specific embodiment, it can also include and the incoherent special knot of analyte on capture pad
Close molecule to a kind of molecule M1, the molecule is connected or combined with the molecule Y1 of specific bond analyte, in detecting pad
Be fixed with detection zone with the incoherent specific binding molecule of analyte to another molecule M2, in label pad wrap
Include mark substance L and another molecule Y1 (Fig. 6 A) with specific bond analyte.When being reacted, if sample
Contain analyte A in this, then the specific binding molecule Y1 on analyte A and capture pad carries out specific bond and forms multiple
Compound matter A-Y1-M1；When the compound substance is reached in label pad by sample pad, new compound substance M1-Y1-A- is just formed
Y2-L；When new compound substance is flowed through on detecting pad, it is fixed on incoherent with analyte in detection zone
Specific binding molecule to the new compound substance of another molecule M2 specific bonds, M2-M1-Y1-A-Y2-L is formed, so that in inspection
There is color on survey region and represent positive findings (Fig. 6 B).
In another specific embodiment, based on competition law, include on capture pad incoherent with analyte
Specific binding molecule to a kind of molecule M1, the molecule is connected or combines with the molecule Y1 of specific bond analyte, in inspection
Survey pad detection zone on be fixed with the incoherent specific binding molecule of analyte to another molecule M2, mark
Include the similar substance A* (Fig. 7 A) of mark substance L and analyte on pad.Reagent concentration on capture pad and test strips can
Arbitrarily adjusted with different with the requirement of detection；, it is necessary to make the concentration of certain drugs in sample big for example in illicit drugs inspection
When the concentration C pre-set, allow colored line is occurred without in detection zone represent positive findings, less than concentration C
When, in detection zone colored line occur represents negative findings.
When being reacted, if contain analyte A in sample, and more than the concentration C pre-set
When, then the specific binding molecule Y1 on capture pad almost carries out specific bond and forms compound substance with analyte A completely
A-Y1-M1, is now likely present the analyte A of remaining excess；When the compound substance by sample pad reaches label pad
On, because the reagent Y1-M1 on capture pad is reacted completely, the reagent A *-L in label pad cannot be attached on Y1-M1；When
When compound substance A-Y1-M1 is flowed through on detecting pad, be fixed in detection zone with the incoherent spy of analyte
Different binding molecule to another molecule M2 specific bonds compound substances, M2-M1-Y1-A is formed, so that in detection zone
Occur without color and represent positive findings (Fig. 7 B).
If on the contrary, containing analyte A in sample, and when less than the concentration C pre-set, then capturing pad
On specific binding molecule Y1 be part and analyte A carry out specific bond formation compound substance A-Y1-M1, now
It is likely present the Y1-M1 of remaining excess；When the compound substance by sample pad reach label pad on, the reagent in label pad
A*-L is just and Y1-M1 combines to form M1-Y1-A*-L；When these compound substances A-Y1-M1, M1-Y1-A*-L are flowed through on detecting pad
When, be fixed in detection zone with the incoherent specific binding molecule of analyte to another molecule M2 it is special
With reference to the compound substance, M2-M1-Y1-A and M2-Y1-A*-L is formed, so that occurring color in detection zone represents negative knot
Really (Fig. 7 B).
The incoherent specific binding molecule of analyte include but not limited to this to (M1/M2), biotin/affinity
Plain (biotin/avidin), biotin/Streptavidin (biotin/streptavidin), antibody/antigen (antibody/
Antigen) (antibody and analyte not including anti-analyte are in itself), the antibody of rhodamine/rhodamine
(rhodamine/anti-rhodamine), mouse IgG/ mouse IgG antibody (Mouse IgG/anti-mouse IgG) etc..
Analyte A can be antibody or antigen, Y1 and Y2 can be on analyte the corresponding specific antigen of different loci or
Antibody molecule or antibody fragment.Mark substance L includes but are not limited to this, colloid gold particle, latex particle, water-soluble
Mark substance etc., what mark substance was connected or combined with some special antigens, antibody or other specific binding molecules
Technology is existing known technology.Certainly, other of some regulation reaction system conditions can also be included in sample application zone
Chemical reagent, such as phosphoric acid buffer reagent, borate buffer reagent, carbonic acid buffer reagent or other albumen, macromolecular etc. come
Optimizing reaction system.These Optimized Measures are all that persons skilled in the art combine the present invention and prior art is readily apparent that
With implementation.
The device in example is applied using upper mask body can detect low concentration or small-molecule substance in sample, because some
The molecular weight very little at the concentration of important analyte in the sample very bottom or analyte, utilizes traditional detection
Device is often because the analyte concentration in sample is very low and can not detect or reach the requirement on clinical meaning.Utilize
The device of the present invention is in advance needing and agent treatment that sample reacts is on capture pad, and one is that can allow sample and reagent reacting
Fully, will not be because the degree that liquid to flow in time and influences reaction to complete, another is exactly that operator need not be still further
The other reagents of addition.
But in actual detection, the above-mentioned detection means for including capture pad is directed to some especially sticky or water content
Sample more than low foam, such as sticky saliva sample, the problem of still occurring following：Because the viscosity of sample is higher, contain
Water is few, or show bubble state, can not sufficiently be reacted with the reagent on capture pad when causing sample on capture pad,
Cause the reagent (antibody or the molecule of specific bond) captured on pad can not be completely combined and as sample flows into the examination in downstream
On paper slip, then in the detection process of test strips, the antigen binding in antibody and label pad without sufficient amount, nothing can be caused
Method develops the color, so as to cause false positive (product detected in particular for competition law).
This problem is solved, then the reagent for needing to enable the analyte in sample fully to be padded with capture is (special
With reference to analyte molecule Y1 and with the incoherent specific binding molecule of analyte to a kind of molecule M1) combine,
One preferred embodiment in, increase by one capture pad (cushion pad), make capture pad include 2, and 2 capture pad separate
It is not connected with, processing has the reagent (Y1, M1) combined with analyte in sample, the examination respectively on two separated capture pads
The total amount of agent is identical with the total amount of an original capture pad.So, sample is made to firstly flow through the first capture pad (capture pad), with catching
Obtain the reagent on pad to carry out after sufficiently reacting, pass through the second capture pad (cushion pad), reacted again with reagent thereon.
So, in the case of reagent total amount identical, the time and space that analyte and reagent are combined all increase, so that
The combination of the two is more abundant, therefore, in the detection process of follow-up test strips, it is to avoid the antibody carried by analyte
Deficiency and the false positive that cause of can not being developed the color in detection zone.
In some specific embodiments, the first capture pad (capture pad) 310 is identical with original capture pulvilliform shape, and difference is
Reagent (Y1, M1 etc.) amount handled thereon is reduced；And the first capture pad (capture pad) 310 is still located on the of detection means
On one collecting chamber 401.Second capture pad (cushion pad) 320 is located in test strips 100, more specifically, positioned at the sample of test strips
On this pad 101, overlapped with the part of sample pad 101, as shown in Figure 4；Processing has another part on second capture pad (cushion pad) 320
Reagent (Y1, M1), sample first flows through the second capture pad (cushion pad) and is combined with reagent thereon, then flowed successively again
Through the sample pad in test strips, label pad, detecting pad and adsorptive pads, detection is completed.
In some specific embodiments, on capture pad (cushion pad) 320 of the first capture pad (capture pad) 310 and second
Reagent content there is certain proportioning, in some specific embodiments, the reagent on the second capture pad (cushion pad) is (special
With reference to analyte molecule Y1 and with the incoherent specific binding molecule of analyte to a kind of molecule M1) in reagent
Between the 15-50% of total amount.Under this proportioning, the combination of analyte and reagent in sample is more abundant.Another is more
For in preferred embodiment, reagent on the second capture pad (cushion pad) (the molecule Y1 of specific bond analyte and with quilt
Analyze the incoherent specific binding molecule of material to a kind of molecule M1) between the 20-35% of reagent total amount.
In a specific embodiment, the production of test strips for convenience, the second capture pad (cushion pad) can be direct
It is identical with sample pad, it is the prolongation of sample pad.After the part processing of extension has Y1 and M1, that is, the second capture pad
The function of (cushion pad), plays a part of the second capture pad (cushion pad).
The material for capturing pad can be polyester fiber film, glass fibre membrane and can stream material selected from capture mat material matter
Filter paper etc..The material of first capture pad (capture pad) and the second capture pad (cushion pad) can be with identical, can also difference.One
In preferred embodiment, the first capture pad (capture pad) material is polyester fiber film；(cushion pad) material is padded in second capture
Glass fibre membrane.
Detected in saliva and be described in detail how with the presence or absence of drugs come exemplified by using the present invention with saliva is collected below
Detection means and detection box.As shown in figure 8, being the collection phase before detection, the first collecting chamber 4011 is located at the second collecting chamber
Inside 4012, and second liquid through hole on the second collecting chamber be not imported with the sample on sample ingress area 406 it is logical
Hole 406-1 is communicated, but is sealed by the baseplane of sample ingress area 406, in order to increase collecting chamber 401 and sample ingress area
The sealing of 306 baseplane, can install sealing in the second liquid through hole correspondence position of the bottom of the second collecting chamber 4012
Circle.The saliva to be detected of collection, first collects the saliva sample for needing to detect, collection first 502 with sample collection device 500 first
It is put into the mouth of detected person, 502 are made up of absorbent material, for example sponge material, so collection head can absorb needs
The saliva sample of detection.Then the first collecting chambers 4011 collection first 502 that suction has saliva sample being put into collecting chamber 401
In, collection first 502 is extruded by rotating the handle 501 of collection device, the saliva sample at this time collected on head is just extruded
Into the first collecting chamber 4011, and by the first liquid through-hole 4011-1,4011-2 is flowed into the second collecting chamber 4012.At this
During individual, because the first capture pad (capture pad) 310 in the first collecting chamber 4011 is prolonged by the first liquid through-hole 4011-1
The bottom of the second collecting chamber 4012 is reached, while sample is extruded, saliva sample is just and on the first capture pad (capture pad) 310
Reagent fully react, after reacting suitable time, such as 10,20 or 30 seconds, 1,2,3,4,5,6,7,8,9,10,15,
20th, 30,45, after 60 minutes, collecting chamber 401 is rotated, is allowed on the second liquid through hole and liquid ingress area of the second collecting chamber
Liquid imports through hole and communicated, and second liquid through hole and liquid import through hole and communicated.This when, saliva sample is just led by liquid
Enter through hole to flow on reagent strip, liquid arrive first at test strips upstream end second capture pad (cushion pad) place, liquid sample after
Continuous to pad the reagent on (cushion pad) with the second capture and sufficiently combined, then, liquid is flowed into sample pad again, successively to
Downstream flow, reaches label pad and detecting pad, finally reaches adsorptive pads, detection reaction is carried out, if existed in saliva a certain amount of
Drugs, colored line is occurred without in the detection zone of reagent strip and represents positive findings, colored line is conversely occurred as soon as and represents
Negative findings, this testing result in detection zone can observe result by the reading window 4041 on body, read
Can be that transparent material covering detection zone can also directly allow detection zone exposed outside on window 4041.Saliva sample
Through hole can also be imported by liquid and enters confirmation chamber 407, saliva sample is detected in order to further confirm.Confirm chamber
407 are surrounded by the side wall of base 4071 and surrounding, can be taken out into the liquid for confirming chamber by exporting through hole 4073, and export is logical
Hole 4073 is sealed before sampling by a plug 4072.When thinking the further by other means of testing result needs, for example, use liquid
Phase color is general, gas chromatography or other more accurate instruments are confirmed when testing result, removes plug 4073, from confirming chamber
407 take out part saliva samples to detect (such as Fig. 8 and 9).Here be only with saliva sample as an example come specifically
It is bright how to use detection means of the invention, in addition to saliva, it can also be other liquid samples, such as blood, urine, sweat
Liquid, excrement etc..
Embodiment
Illustrate how the present invention realizes that illustrated now with limited experiment, these explanations only exist in order to clearer
Limited citing is done under the marrow of the claim of the present invention, limiting to the claimed invention is not constituted.
Experiment：With the present invention improve before and after detection means detection saliva in the presence or absence of certain density anxiolytic with
Hypnotic sedative agent (BZO).
Part I：The reagent strip of the present invention and the assembling of detection means and part, which make, (has the first capture pad (capture
Pad) and the second capture pad (cushion pad))
Reagent strip 100 as shown in Figure 3 is used for illustrating the part and preparation method of the reagent strip of this experiment present invention used.
The processing of nitrocellulose filter.Detecting pad 103 is nitrocellulose filter (NC), there is two on nitrocellulose filter
Lines, one is detection line 132, the testing result control line 133 positioned at detection line downstream.It is fixedly connected with detection line
IgG Streptavidin (streptavidin-IgG)；The fixed foster anti-rabbit IgG on testing result control line.Fixed method
It can be automatically processed with automatic spray film process machine, wherein the concentration in processing detection lines is 1.0 mg/mls, dilution
Buffering liquid is phosphate buffer (PBS)；Goat anti-rabbit igg antibody is fixed on testing result control line；Concentration be 1.0 milligrams/
Milliliter, dilution buffer liquid is phosphate buffer (PBS).The nitrocellulose filter handled well is placed in 37 DEG C of baking ovens and dried i.e.
Can.
The processing of label pad 102.Marker slip is polymer PET, divides BSA with coupled protein by what colloid gold particle had been marked
BZO haptens be processed on polymer PET.The OD values of processing solution are 57, and dilute solution is 1 times of PBS cushioning liquid, wherein
Also containing 1% BSA.Also processing has the rabbit igg antibody of colloid gold label on marker slip.The marker slip handled well is placed on 37
Dried in DEG C baking oven.
The processing of sample pad 101.Sample blank film is the plain film of glass fibre, and the solution handled on the film is：Borax
(0.07M/L)；Tween20 (1%)；Cholic Acid (1%)；Tris(0.1M).The sample blank film handled well is placed on 37
Dried in DEG C baking oven.
The processing of first capture pad (capture pad) 310.Capture pad includes paste-side 3101 and the water imbibition of unwetted
Reagent end 3102.The material at reagent end is polyester film, and the chemical solution of reagent end processing includes：The upper biotin of connection
(Biotin) anti-BZO antibody materials are dissolved in 1 times of PBS cushioning liquid, and it is 1.0 mg/mls to make ultimate density；Wherein
Also containing 1% BSA, it is 0.0025 milligram to make the handle well first anti-BZO antibody materials content captured on pad (capture pad).
The capture pad handled well is placed in 37 DEG C of baking ovens and dried.Then the polymer PET small pieces (3102) containing reagent are pasted onto
On the small pieces of unwetted on (3101).
The processing of second capture pad (cushion pad) 320.The capture pad of a part second (cushion pad) is located on whole capture pad
Physicochemical solution：The anti-BZO antibody materials of the upper biotin (Biotin) of connection are dissolved in 1 times of PBS cushioning liquid, made most
Final concentration of 0.2 mg/ml；Wherein also containing 1% BSA；Handle well second is set to capture the anti-BZO padded on (cushion pad)
Antibody materials content is 0.0005 milligram.The capture pad of another part second (cushion pad), by 0.2 milligram/milli in the way of a spray
On the second capture pad position of (cushion pad) away from the linear series at a port 2mm that the anti-BZO antibody materials risen are sprayed on.Will
The second all capture pads (cushion pad), which is placed in 37 DEG C of baking ovens, dries.
Content ratio in the final anti-BZO antibody materials content made on first sample pad and the second capture pad (cushion pad) is about
For 5：1.
The assembling of reagent strip 300.The all parts handled well are assembled according to the appearance shown in Fig. 4, the second capture pad is allowed
(cushion pad) (two kinds) upstreams positioned at sample pad, allow sample pad to be located at the upstream of label pad, and label pad is located at detecting pad and sample
Between this pad, the filter paper 104 of water suction is placed in the downstream of detecting pad.These parts are all placed on the support chip of a unwetted
On 105.
The assembling of detection means.Detection means such as Fig. 8, shown in 9.Glue the paste-side 3101 that (capture pad) is padded in first capture
On the surface for the vertical bar beam being attached on the first collecting chamber 4011, the other end 3102 is through liquid through-hole and reaches the second collecting chamber
4012 bottom.Then whole collecting chamber 401 is arranged in the snap ring of the sample Lead-In Area 406 of body 404, and allows second
The bottom of collecting chamber 4012 is in the position for the basal surface sealing (Fig. 8) for being imported into region.The reagent strip 300 with capture pad
It is placed between upper plate 4043 and lower plate 4045, makes the detection zone of reagent strip 100 corresponding with reading window 4041, sample receives
It is corresponding that region imports through hole 306-1 with liquid.Upper plate and lower plate, which are fastened, to be integral.Thus it has been assembled into used in this experiment
Detection means.
Part II：The assembling and making of control stripes bar 100 and detection means 400 (there is capture pad).
Compared with reagent strip recited above and detection means, contrast agents bar and detection means are substantially the same.With
Unlike, install on detection means for capture pad, while in test strips without second capture pad (cushion pad).
Wherein, the processing of pad 210 is captured.The chemical solution of the reagent end processing of capture pad includes：The upper biotin (Biotin) of connection
Anti- BZO antibody materials be dissolved in 1 times of PBS cushioning liquid, make ultimate density be 1.0 mg/mls；Wherein also contain
1% BSA, it is about 0.003 milligram to make the anti-BZO antibody materials content on the capture pad handled well.
Testing result record is all read after saliva sample is added 10 minutes.Generally, set the concentration of BZO in saliva sample as
10ng/ml (nanograms/milliliter) is detection threshold value.If detection threshold value it is meant that analyte BZO in sample
Concentration is more than detection threshold value, in theory, and testing result should be positive, whereas if analyte BZO concentration is less than
Detection threshold value, testing result is feminine gender.
Operating method：
The bottom for first allowing each detection means to be in the second collecting chamber 4012 of collecting chamber 401 is imported into region 406 and closed
State.
Then the sample to be detected is added, is allowed on capture pad in sample and collecting chamber 401 (the first capture pad (capture pad))
Reagent reacting 1 minute；
Rotating collection chamber 401, allows saliva fluid to flow to the sample application zone of reagent strip from collecting chamber and completes reaction；
The actual result of detection is recorded after 10 minutes.
Explanation：
Control is the detection means before improving；Lot1 is device after improving, wherein the second capture pad (cushion pad) is overall
Reagent treatment；Lot2 is device after improving, wherein second capture pad (cushion pad) linear process reagent
1. compare the detection sensitivity of detection means before and after improving using standard solution.Set respectively in master sample
The concentration for putting BZO is 0ng/ml (negative sample)；30ng/ml, is detected using each device.
Analysis：
Detection means after improvement is preceding higher than improving in detection sensitivity.
2. the detection of pair negative sample：The saliva sample of collection our company employee is detected
Analysis：
False positive sample before improvement, after being detected using the detection means after improvement, is completely eliminated.Therefore, should
Detection means after improvement can avoid the presence of the false positive of detection, improve the accuracy rate of detection.
3. the detection to positive sample：
Multiple negative saliva samples are gathered, the BZO (seeing attached list) of various concentration is separately added into these salivas, is entered respectively
Row detection
Analysis：
In terms of upper table, the detection means after improvement, when detecting positive sample, changes compared with before improvement without any
Become.Therefore, the detection means after improvement does not have any influence when detecting positive sample.
In summary analysis of experimental results：
1. the detection means after improving increases compared with before-improvement in detection sensitivity；
2. the detection means after improving can effectively avoid the false positive issue of negative sample, the accurate of detection is improved
Rate；
3. the detection means after improving is identical with before improvement when detecting positive sample, do not influence what positive sample was detected
Accuracy.
Claims (12)
1. a kind of sample testing apparatus, including test strips, test strips include sample pad, label pad and detecting pad, label pad is located at
The downstream of sample pad, detecting pad is located at the downstream of label pad, it is characterised in that the sample testing apparatus also includes collecting chamber, caught
Pad and body are obtained, capture pad is located in collecting chamber, and test strips are located within the body, the test paper in capture pad and body in collecting chamber
Bar liquid communication, and capture pad is positioned at the upstream of test strips fluid course.
2. sample testing apparatus according to claim 1, it is characterised in that sample testing apparatus also includes cushion pad, delays
Punching pad is connected positioned at the upstream of the sample pad of test strips with sample pad.
3. sample testing apparatus according to claim 2, it is characterised in that the body also includes sample ingress area,
Collecting chamber is connected with ingress area；Have on ingress area and import through hole, the importing through hole makes ingress area and body pilot scale
Paper slip liquid communication.
4. sample testing apparatus according to claim 3, it is characterised in that collecting chamber includes the first collecting chamber and second and received
Collect chamber, the first collecting chamber is in the second collecting chamber；First collecting chamber, which has, makes the first collecting chamber be connected with the second collecting chamber liquid
The first logical liquid through-hole；There is second liquid through hole on second collecting chamber.
5. sample testing apparatus according to claim 4, it is characterised in that collecting chamber can turn relative to ingress area
It is dynamic；Before rotation, the second liquid through hole of the second collecting chamber is imported into the baseplane sealing in region, collecting chamber bottom is imported into area
Domain bottom surface is sealed；After rotation, the second liquid through hole of the second collecting chamber is connected with the importing through hole on ingress area, makes receipts
Collect chamber and ingress area liquid communication.
6. sample testing apparatus according to claim 4, it is characterised in that capture pad is loaded with reagent, capture pad is being received
Collection intracavitary position cause liquid by the first collecting chamber flow into the second collecting chamber during, or liquid be located at first or second
During collecting chamber, it can be in contact with the reagent on capture pad.
7. sample testing apparatus according to claim 6, it is characterised in that capture pad and the first collecting chamber side wall or bottom wall
It is connected, either capture pad is located in the second collecting chamber or the first collecting chamber is run through in capture pad one end, its one end is located at first
Collecting chamber, the other end is located at the second collecting chamber；Or capture pad is formed positioned at the first collecting chamber bottom with the second collecting chamber bottom
Insulating space in.
8. the sample testing apparatus according to one of claim 1-7, it is characterised in that cushion pad is the extension of sample pad
Point.
9. the sample testing apparatus according to one of claim 1-7, it is characterised in that capture mat material matter is polyester fiber
Film；Cushion pad material is glass fibre membrane.
10. the sample testing apparatus according to one of claim 1-7, it is characterised in that included on capture pad and cushion pad
The molecule of specific bond analyte and with the incoherent specific binding molecule of analyte to a kind of molecule.
11. sample testing apparatus according to claim 10, it is characterised in that the specific bond analyte on cushion pad
Molecule and with the incoherent specific binding molecule of analyte to a kind of molecule content for capture pad and cushion pad
The 15%-50% of total amount.
12. sample testing apparatus according to claim 10, it is characterised in that the molecule of specific bond analyte and
With the incoherent specific binding molecule of analyte to a kind of molecule be processed on whole capture pad；Specific bond quilt
The molecule of analyte and with the incoherent specific binding molecule of analyte to a kind of molecule in linearly aligned mode
It is processed on a line of cushion pad.
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| CN105223349A (en) | 2016-01-06 |
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