CN103336135A - Procalcitonin detection test strip - Google Patents
Procalcitonin detection test strip Download PDFInfo
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- CN103336135A CN103336135A CN201310257890XA CN201310257890A CN103336135A CN 103336135 A CN103336135 A CN 103336135A CN 201310257890X A CN201310257890X A CN 201310257890XA CN 201310257890 A CN201310257890 A CN 201310257890A CN 103336135 A CN103336135 A CN 103336135A
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- procalcitonin
- region
- test strip
- ucp
- monoclonal antibody
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Abstract
The invention relates to the field of biotechnology, and discloses a procalcitonin detection test strip. The procalcitonin detection test strip comprises a sample loading region, a conjugate release region, a reaction region and an absorbent pad which are orderly arranged; the conjugate release region is a conjugate release pad of an up-converting phosphor (UCP) marked procalcitonin monoclonal antibody; the reaction region is divided into a testing region and a control region, and the reaction region is a solid support in which the testing region coats procalcitonin and the control region coats a Goat anti Mouse IgG antibody. The procalcitonin detection test strip can quickly, simply and quantitatively detect the content of a detected object, and provide supports for diagnosis of bacterial infection, differential diagnosis, therapeutic effect monitoring and estimation after recovery.
Description
Technical field
The present invention relates to biological technical field, be specifically related to a kind of Procalcitonin test strip.
Background technology
(procalcitonin PCT) is the calcitonin precursor substance of no hormonal activity to Procalcitonin, is made up of 116 amino acid, and molecular weight is the glycoprotein of 13KD.PCT secretes (comprising thyroid gland, lung and pancreatic tissue cell) by neural inner cell and expresses, and cuts through enzyme and is decomposed into calcitonin (CT), c-terminal peptides and amino terminal peptide.The half life period of PCT in human body is 25-30h, good stability, and content is extremely low in normal human serum.
Under the many situations that cause SIRS (SIRS), such as bacterial infection, pancreatitis, burn to death, multiple injury, the content of PCT raises unusually in the blood, and CT does not have significant change.The order of severity of PCT level and infection and damage is proportionate in the blood, and has become for pyemia, severe sepsis, the diagnosis of septicopyemia shock and the efficiency index of curative effect monitoring.
Some organic or inorganic materials that occurring in nature exists can be excited by electromagnetic radiation, and emitting fluorescence or phosphorescence are all observed the Stokes rule in its luminous process, and namely radiative wavelength is longer than and is excited light wavelength.The seventies in 20th century, scientist has found that a class can anti-Stokes rule produces the special material of phosphorescence, this material is at infrared light district (wavelength＞780nm) be excited, but can the long-range visible light (wavelength 475nm-670nm) that is shorter than exciting light of transmitted wave, be to change on the energy, this phenomenon is called as transmits light.
Be doped in the material that constitutes in the lattice of crystal by some thulium and have the forwarding of going up optical phenomenon, three kinds of major ingredients are arranged: main matrix (host matrix), absorption (absorber) and emission (emitter) in this material.Crystalline material as main matrix has: oxysulfide is (as Y
2O
2S, GdO
2S, La
2O
2S etc.), fluoride is (as YF
3, GdF
3, LaF
3Deng), sow hydrochlorate (as YGaO
3, Y
3Ga
5O
12Deng) and silicate (as YSi
2O
5, YSi
3O
7Deng) etc.;
Being commonly used for the rare earth ion that absorbs son has: ytterbium ion (Yb
3+), erbium ion (Er
+), samarium ion (Sm
3+) etc.; The rare earth ion that is commonly used for emission has: erbium ion (Er
+), holmium ion (Ho
3+), thulium ion (Tm
3+), terbium ion (Tb
3+) etc.Spatial orientation and distance that absorption and this ion pair of emission suit in the main matrix lattice are to produce to go up the basis of transmitting light.The generation of last forwarding light is a two-phonon process that relates to a plurality of photons (at least two).In this process, on transmit absorption in the luminescent material (as Yb
3+) to absorb two energy photons (infrared light districts at least, as 970nm), then through a series of internal energy conversions, with radiationless form (A1 → A2, A2 → A3) energy with these two photons passes to emission (as Era+) continuously, so that it is in excited state (A3).A transition of returning ground state level then takes place in the latter, discharges a high-energy photon (visible region is as 525nm or 540nm) and finishes on the energy and change.
Transmit the process (see figure 1) of light in two photon excitation UCP generation.
(up-converting particle UCP) has different optical properties in conjunction with making forwarding light particle for different absorption, emission, main matrix.(1) under the situation that main matrix is identical, a series of UCP adopt identical absorption, different emission, and what then it can be by identical wavelength is infrared ray excited, produces emission light (as: the infrared ray excited absorption-Yb of 980nm of different wave length
3+-emission-Er
3+, absorb son-Yb
3+-emission-Tm
3+, launch the green visible light of 550nm, 475nm blue visible light respectively); Adopt different absorption, identical emission is though then via different exciting lights, can produce identical emission light.(2) under the different situation of main matrix, the UCP spectral signature also will change and (as: absorb son-Yb
3+-emission-Er
3+, at main matrix YF
3, GdF
3In, emission redness respectively and green visible light).The diversity of forming has determined the diversity of UCP spectrum (excitation spectrum, emission spectrum), and this becomes the basis of dirigibility in its use.
The method that detects PCT at present has a variety of, except the past working costs is difficult for the gel layer analytic approach and high efficiency liquid phase chromatographic analysis method of robotization, detecting more special, the responsive method of PCT now has: enzyme linked immunosorbent assay, radioimmunology, immunofluorescence technique and colloidal gold immunity chromatography.
Enzyme linked immunosorbent assay is to utilize two kinds of mouse monoclonal antibody double antibody sandwich method principles to detect PCT in the serum, and method is more special, no cross reaction.But the enzyme linked immunosorbent assay detection sensitivity is low, the minimum only 10ug/L that is limited to, and PCT concentration can not detect in the normal human serum.
Radioimmunology can detect free type PCT, can detect mating type PCT again, detection sensitivity height, the minimum 4pg/L that is limited to.But it is longer that radioimmunology detects required time, need a few hours usually, and the pollution of radioelement limited the application of the method.
Immunofluorescence technique is a kind of quantitative immunofluorescence detection method, two kinds of binding site combinations that mouse monoclonal antibody is different with two of PCT antigen, a kind of antibody is through fluorescence labeling (tracer agent), another kind is fixed on test tube wall, PCT molecular reaction in antibody and the serum forms " sandwich complex ", fluorescent-labeled antibody is tied up on test tube wall, content by luminescence reagent numeration fluorescent marker, fluorescence signal intensity is directly proportional with sample PCT concentration, while bioassay standard product, after making typical curve according to the PCT of known antigens concentration, quantitatively to obtain the PCT concentration of sample, the method susceptibility is similar to radioimmunology.But immunofluorescence technique need be equipped with expensive immunofluorescence detector.
Colloidal gold immunity chromatography mainly is to use a monoclonal mouse-anti Procalcitonin antibody to be combined in (tracer) and polyclone goat-anti one a Procalcitonin antibody (solid phase) on the collaurum, after patient specimen is added to reacting hole, tracer is in conjunction with the PCT in the sample and form tangible antigen antibody complex, by observing the district, here the antigen antibody complex of mark is attached on the fixing anti-Procalcitonin antibody and forms sandwich complex this compound through syphonic effect.When PCT concentration＞0.5ug/L, the sandwich complex band that takes on a red color, the PCT concentration in shade and the sample is in direct ratio, unconjugated tracer is diffused into the check plot, in conjunction with and produced red contrast band.Colloidal gold immunity chromatography is simple fast, but only can half-quantitative detection.
Summary of the invention
In view of this, the invention provides a kind of fast, accurately, the former test strip of the Procalcitonin in the detection human serum that can be quantitative and preparation method thereof.
In order to realize the foregoing invention purpose, the invention provides following technical scheme:
A kind of Procalcitonin test strip comprises that the sample application zone, the bond that set gradually discharge district, reaction zone and adsorptive pads; Described bond discharge the district for bag by the last forwarding luminescent material Procalcitonin monoclonal antibody that is the UCP mark; Described reaction zone is for being divided into test section (T district) and control zone (C district), and described reaction zone is that wrap by the Procalcitonin monoclonal antibody test section, and wrap by the solid support of sheep anti-mouse igg antibody the control zone.
Procalcitonin in the quantitative test sample of Procalcitonin quick detection test paper strip adoption double-antibody sandwich immune chromatography method.Wrap in advance by anti-Procalcitonin monoclonal antibody test section on the analyzing film (T), and Quality Control district (C) wraps in advance by sheep anti-mouse igg, and bond discharges the anti-Procalcitonin monoclonal antibody of another strain that UCP particle mark is arranged in the district.Procalcitonin and two strain antibody reaction bonded in the sample during reaction, form the Procalcitonin protein antibodies compound of Procalcitonin protein antibodies-Procalcitonin proteantigen-UCP mark in the T district, the UCP particle is at the infrared ray excited visible light of emission down, and radiative intensity is directly proportional with the concentration of Procalcitonin albumen in the sample.No matter whether there is Procalcitonin in the sample, all can forms the anti-Procalcitonin antibody complex of sheep anti mouse-UCP mark in the C district.Utilize the UPT biology sensor that test strips is carried out scanning analysis, show measurement result.
Control zone (C district) bag is by sheep anti-mouse igg antibody, on to transmit luminescent material be that the sheep anti-mouse igg reaction back scanning analysis of Procalcitonin monoclonal antibody and the bag quilt of UCP mark does not have the result, illustrate that then test strips lost efficacy.
Further, Procalcitonin test strip of the present invention also comprises backing, and what described sample application zone, bond release district, reaction zone and adsorptive pads overlapped successively mutually sticks on the backing.
Because the concentration of Procalcitonin is that the intensity of UCP decides by transmitting luminescent material on the amplification of signal thing that is excited finally, so the Procalcitonin monoclonal antibody has been played the part of, and the forwarding luminescent material is the key player of UCP and these two kinds of materials of Procalcitonin in the connection, can be that UCP is combined with last forwarding luminescent material, can be combined with Procalcitonin again, wherein to be combined with Procalcitonin be a spontaneous process to the Procalcitonin monoclonal antibody, and Procalcitonin monoclonal antibody and last forwarding luminescent material be UCP in conjunction with needing artificial promotion.An important step of producing goes up exactly and transmits luminescent material is UCP mark Procalcitonin monoclonal antibody.Mark the last forwarding luminescent material Procalcitonin monoclonal antibody that is UCP must stable performance, be dissolved in do not have precipitation after the damping fluid, not having free going up, to transmit luminescent material be UCP.
Wherein, as preferably, the Procalcitonin MONOCLONAL ANTIBODIES SPECIFIC FOR method that upward forwarding luminescent material of the present invention is the UCP mark is to 1mL pH7.4, adding the last forwarding of 2umol luminescent material in the phosphate buffer of 50mM is the former monoclonal antibody of the anti-HCT of UCP and 1mg, gentle vibration is 2 hours under the room temperature, the centrifugal 30min of 12000r/min gets precipitation namely then.
Wherein, described Procalcitonin monoclonal antibody, Procalcitonin and sheep anti-mouse igg antibody are well known to a person skilled in the art, can obtain or prepare by disclosed method in the prior art by commercial sources.
Transmitting luminescent material as going up of amplification of signal thing is that UCP is the emphasis during test strips quantitatively detects.Last forwarding luminescent material be the size of UCP, all once, stability all can influence the last reading of product.As preferably, in the Procalcitonin test strip of the present invention, describedly go up that to transmit luminescent material be that the UCP particle diameter is 5nm.
As preferably, described bond discharges pad and is the plain film of glass fibre.
In the present invention, the solid support of reaction zone is nitrocellulose filter.
Sample application zone of the present invention and adsorptive pads are the material with absorption function, are preferably thieving paper.
It is fixing that backing of the present invention supports that sample application zone, bond discharge district, reaction zone and adsorptive pads bonding.Described backing is preferably the PVC plate.
Procalcitonin of the present invention detects test paper and comprises that the sample application zone, the bond that set gradually discharge district, reaction zone and adsorptive pads; Described bond discharges the Procalcitonin monoclonal antibody that the district is the UCP mark for bag by last forwarding luminescent material; Described reaction zone is for being divided into test section (T district) and control zone (C district), and described reaction zone is that wrap by the Procalcitonin monoclonal antibody test section, and wrap by the solid support of sheep anti-mouse igg antibody the control zone.The above luminescent material of transmitting of Procalcitonin test strip of the present invention is that UCP substitutes traditional collaurum, and the forwarding luminescent material is the Procalcitonin in the UCP immunochromatographyassay assay human serum in the utilization.Compare with traditional colloidal gold method, test strips of the present invention not only sensitivity improves tens of times, but also can quantitatively detect, for clinical diagnosis and treatment provide foundation more accurately.The large-scale expensive instrument of the required instrument of this test strips clinical laboratory different from the past, only need a small-sized UPT biology sensor (on transmit the light immunity analysis instrument) to get final product, realized real-time detection, for hospitals at different levels and Disease Control and Prevention Center provide detection means easily and fast to Procalcitonin quantitative Diagnosis and epidemiology survey.
Description of drawings
Fig. 1 shows the side schematic view of Procalcitonin test strip of the present invention, wherein 1: and sample application zone, 2: bond discharges district, 3: reaction zone, 4: adsorptive pads, 5: backing;
Fig. 2 shows the front schematic view of Procalcitonin test strip of the present invention, and 1: sample application zone, 2: bond discharges district, 4: adsorptive pads, T: test section, C: control zone.
Embodiment
The invention discloses a kind of Procalcitonin test strip.
The used biomaterial of a kind of Procalcitonin test strip provided by the invention and reagent all can be buied by market.
Below in conjunction with embodiment, further set forth the present invention:
Embodiment 1: bond discharges the preparation in district to 1mL pH7.4, adding the 2umol particle diameter in the phosphate buffer of 50mM is 5nm, excitation wavelength is 365nm, emission wavelength is that the last forwarding of 605nm luminescent material is UCP and the former monoclonal antibody of 1mg HCT, gentle vibration is 2 hours under the room temperature, the centrifugal 30min of 12000r/min then gets precipitation and gets namely that to transmit luminescent material be the Procalcitonin monoclonal antibody of UCP mark.
The last forwarding luminescent material Procalcitonin monoclonal antibody that is the UCP mark after stabilizing agent is handled, evenly is sprayed on the plain film of glass fibre by the amount of every square centimeter of 65uL, and freeze drying namely gets bond and discharges the district.
Embodiment 2: the preparation of reaction zone
1mg/mL Procalcitonin 10uL is sprayed on the NC Nitroncellulose film of reaction zone, puts freeze drying and namely get the test section.
1mg/mL sheep anti-mouse igg antibody 10uL is sprayed on the NC Nitroncellulose film of reaction zone, puts freeze drying and namely get the control zone.
Embodiment 3: the preparation of test strips
The bond that sample application zone, embodiment 1 are made discharges reaction zone that district, embodiment 2 make and adsorptive pads sticking on the backing namely of overlap joint mutually successively.
The above only is preferred implementation of the present invention; should be pointed out that for those skilled in the art, under the prerequisite that does not break away from the principle of the invention; can also make some improvements and modifications, these improvements and modifications also should be considered as protection scope of the present invention.
Claims (5)
1. immuno-chromatographic test paper strip based on up-converting phosphor technology is characterized in that: Procalcitonin monoclonal antibody of having transmitted light UCP particle mark on it.
2. the described immuno-chromatographic test paper strip of claim 1, it is characterized in that: reaction zone is nitrocellulose filter, and the test section on the reaction zone is coated with the Procalcitonin monoclonal antibody, and bag is by sheep anti-mouse igg antibody on the control zone.
3. the described immuno-chromatographic test paper strip of claim 1 is characterized in that: by described well, biological sample is added on the sample pad; The interpretation window is corresponding to the test section on the reaction zone and control zone as a result; The terminal point indication window is corresponding to the terminal point index strip on the adsorptive pads.
4. the described immuno-chromatographic test paper strip of claim 1 is characterized in that: described bond discharges the district and is the plain film of glass fibre.
5. test sample is blood sample according to claim 1.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310257890XA CN103336135A (en) | 2013-06-20 | 2013-06-20 | Procalcitonin detection test strip |
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310257890XA CN103336135A (en) | 2013-06-20 | 2013-06-20 | Procalcitonin detection test strip |
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| CN103336135A true CN103336135A (en) | 2013-10-02 |
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Citations (8)
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| US20040121343A1 (en) * | 2002-12-24 | 2004-06-24 | Biosite Incorporated | Markers for differential diagnosis and methods of use thereof |
| CN2706765Y (en) * | 2004-04-23 | 2005-06-29 | 中国人民解放军军事医学科学院微生物流行病研究所 | Immunochromatography test paper strip based on up-conversion luminescence technology |
| CN102116770A (en) * | 2009-12-31 | 2011-07-06 | 北京热景生物技术有限公司 | Immunochromatography rapid kit and production method thereof |
| CN102636642A (en) * | 2012-04-06 | 2012-08-15 | 中国人民解放军第三0二医院 | Quick quantitative kit for hepatic fibrosis diagnosis |
| CN202548130U (en) * | 2012-04-06 | 2012-11-21 | 中国人民解放军第302医院 | Improved immunochromatography test paper strip |
| CN202794178U (en) * | 2012-08-07 | 2013-03-13 | 天津中新科炬生物制药有限公司 | Fast quantitative immunochromatographic assay kit for procalcitonin |
| CN202814980U (en) * | 2012-08-09 | 2013-03-20 | 河南省农业科学院 | Immuno-chromatographic test strip for quantitative detection of clenbuterol based on up-conversion fluorescence nano-particle marks |
| CN103018456A (en) * | 2012-11-29 | 2013-04-03 | 深圳市伯劳特生物制品有限公司 | Procalcitonin detection test strip and preparation method thereof |
-
2013
- 2013-06-20 CN CN201310257890XA patent/CN103336135A/en active Pending
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20040121343A1 (en) * | 2002-12-24 | 2004-06-24 | Biosite Incorporated | Markers for differential diagnosis and methods of use thereof |
| CN2706765Y (en) * | 2004-04-23 | 2005-06-29 | 中国人民解放军军事医学科学院微生物流行病研究所 | Immunochromatography test paper strip based on up-conversion luminescence technology |
| CN102116770A (en) * | 2009-12-31 | 2011-07-06 | 北京热景生物技术有限公司 | Immunochromatography rapid kit and production method thereof |
| CN102636642A (en) * | 2012-04-06 | 2012-08-15 | 中国人民解放军第三0二医院 | Quick quantitative kit for hepatic fibrosis diagnosis |
| CN202548130U (en) * | 2012-04-06 | 2012-11-21 | 中国人民解放军第302医院 | Improved immunochromatography test paper strip |
| CN202794178U (en) * | 2012-08-07 | 2013-03-13 | 天津中新科炬生物制药有限公司 | Fast quantitative immunochromatographic assay kit for procalcitonin |
| CN202814980U (en) * | 2012-08-09 | 2013-03-20 | 河南省农业科学院 | Immuno-chromatographic test strip for quantitative detection of clenbuterol based on up-conversion fluorescence nano-particle marks |
| CN103018456A (en) * | 2012-11-29 | 2013-04-03 | 深圳市伯劳特生物制品有限公司 | Procalcitonin detection test strip and preparation method thereof |
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Application publication date: 20131002 |