CN213210176U - Device for synchronously detecting different types of influenza viruses and pneumonia IgM antibodies - Google Patents
Device for synchronously detecting different types of influenza viruses and pneumonia IgM antibodies Download PDFInfo
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- CN213210176U CN213210176U CN202020915522.5U CN202020915522U CN213210176U CN 213210176 U CN213210176 U CN 213210176U CN 202020915522 U CN202020915522 U CN 202020915522U CN 213210176 U CN213210176 U CN 213210176U
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Abstract
The utility model belongs to the field of medical detection equipment, and provides a device for synchronously detecting different types of influenza viruses and pneumonia IgM antibodies, which comprises a double-channel card shell, test paper strip FluA & B and test paper strip CP & MP which are arranged in the double-channel card shell in parallel; the test strip FluA & B is characterized in that a high-specificity influenza A virus antigen is coated on an influenza A virus detection line, a high-specificity influenza B virus antigen is coated on an influenza B virus detection line and a quality control line coated receptor is coated on a first quality control line which are arranged on a nitrocellulose membrane at intervals; the test paper strip CP & MP is characterized in that a chlamydia pneumoniae detection line (CP) arranged on a nitrocellulose membrane at intervals is coated with a high-specificity chlamydia pneumoniae antigen, a mycoplasma pneumoniae detection line (MP) is coated with a high-specificity mycoplasma pneumoniae antigen, and a second quality control line is coated with a quality control line coating receptor.
Description
Technical Field
The utility model relates to a medical treatment check out test set technical field generally, specifically relate to a device of synchronous detection different grade type influenza virus and pneumonia IgM antibody
Background
Influenza virus type a/B is the major pathogen of influenza and can cause serious complications in patients with underlying pathology. Among them, influenza A antigen is easy to be mutated, causing a worldwide pandemic for many times; influenza type B is less pathogenic to humans and has a much smaller epidemic size than type a. Influenza viruses belong to myxoviruses, and have a polymorphic form, generally spherical, and also filamentous or rod-shaped, with a virus diameter of 80-120 nm. The structure of influenza viruses consists primarily of an inner core, a capsid, and an outer envelope.
The main source of infection with influenza viruses is the patient, and secondly the recessive infector, and infected animals may also be a source of infection. The main transmission route is droplets with influenza virus, which enter the body through the respiratory tract. A few may also be infected by indirect contact with common handkerchiefs, towels, etc.
Chlamydia Pneumoniae (CP) is a newly discovered chlamydia, mainly causing respiratory and pulmonary infections. Chlamydia pneumoniae often produces upper respiratory and respiratory tract infections in children and adults. Only humans are known to be the chlamydial host and the mode of infection may be interpersonal transmission through respiratory secretions. Chlamydia pneumoniae infection in humans is worldwide. The chlamydia pneumoniae infection route is human-to-human transmission through respiratory secretions. Thus, there may be a small range of popularity in semi-enclosed environments such as homes, schools, military and other work areas where populations are concentrated. Infection with chlamydia pneumoniae may also be associated with the onset of asthma, coronary heart disease and atherosclerosis, acute episodes and exacerbations of chronic obstructive pulmonary disease. Currently, chlamydia pneumoniae is the main pathogen causing community-acquired pneumonia after diplococcus pneumoniae and haemophilus influenzae, and becomes three atypical pathogens of community-acquired pneumonia together with legionella pneumophila and mycoplasma pneumoniae, accounting for 10% -20% of community-acquired pneumonia. Children under 5 years old are rarely infected, and children over 8 years old and young are susceptible to infection.
Mycoplasma Pneumonia (MP) is acute respiratory infection with pneumonia caused by Mycoplasma pneumoniae, the diameter of Mycoplasma pneumoniae is 125-150 nm and is between bacteria and viruses, and the Mycoplasma pneumonia is propagated by oral and nasal secretions through air to cause respiratory infection, and is more in autumn and winter. MP is one of the important pathogens of pneumonia and other respiratory infections in childhood. Mycoplasma pneumoniae can cause tonsillitis, rhinitis, otitis media, tracheitis, bronchiolitis and pneumonia, children mostly suffer from upper respiratory tract infection, and school-age children mostly suffer from pneumonia.
At present, the clinical common influenza virus A/B rapid detection mainly takes antigen detection, and nasopharyngeal swab collection is taken as a detection specimen, while the common Chlamydia Pneumoniae (CP) and mycoplasma pneumoniae rapid detection methods mainly take antibody detection, blood samples are taken as detection specimens, and relatively mature detection devices, such as detection test paper, detection kits and the like, are developed, but the detection devices are used for singly detecting the antigen of a virus or a bacterium, such as CN105823875A, and the mycoplasma pneumoniae antibody immune colloidal gold detection test paper and the preparation method thereof are disclosed; or simultaneously detecting two same types of bacteria or viruses, such as CN109142725A discloses a preparation method of the colloidal gold joint detection test paper for the influenza A universal virus and the H7N9 subtype virus.
Since influenza virus, chlamydia, mycoplasma infections are easily confused with other respiratory diseases, the main problem between the existing detection devices and clinical diagnosis is represented by the following aspects:
1. the requirements for detecting samples are not uniform, and multiple tests and diagnoses are required: the current clinical treatment mode is that when influenza virus A/B, Chlamydia Pneumoniae (CP) and mycoplasma pneumoniae are rapidly identified and diagnosed according to clinical symptoms, a laboratory sheet is required to be provided for many times, different detection specimens are required to be collected for many times, and multiple detection reagents of the same manufacturer or different manufacturers are adopted to perform multi-step complex operations.
2. Because of the discontinuities and incompleteness between multiple assays and/or diagnoses, medication errors or prolonged illness: when influenza virus type A/B and Chlamydia Pneumoniae (CP)/mycoplasma pneumoniae are detected singly and clinically judged according to results, the fact that the illness state cannot be comprehensively analyzed and the materials cannot be known occurs, and timely treatment is affected because a patient is reluctant to carry out detection repeatedly.
Therefore, the device and the method which can simultaneously detect and judge or eliminate the acute infection of influenza virus A/B and Chlamydia Pneumoniae (CP)/Mycoplasma pneumoniae by the same sampling have great innovative significance in clinical practice.
In addition, a trend chart of IgM and IgG antibodies after pathogen infection is shown in figure 1, and it can be seen that the latency period of common respiratory tract pathogen infection is 1-7 days, serum specific antibodies are gradually generated 7-10 days after infection, firstly, immunoglobulin IgM antibodies appear, IgM is the antibody appearing at the earliest in the primary humoral immune response, and is the first army of the organism resisting infection, and then the IgG antibodies appear, and IgM appears early and disappears fast, is closely related to the disease course development, and IgG appears late and disappears slowly and lags behind the disease course development stage. Thus, the presence of IgM antibodies suggests a recent infection and the presence of IgG antibodies suggests a previous infection.
Therefore, by simultaneously detecting IgM of four respiratory pathogens in a blood sample and using the IgM in an early diagnosis of respiratory pathogen infection, sensitivity and accuracy are improved compared with other indexes.
Disclosure of Invention
The utility model aims at the defect of prior art and the urgent need of clinical, provide a synchronous detection influenza virus A type, B type and pneumonia chlamydia, the IgM antibody's of mycoplasma coalition unit, the utility model discloses the device passes through the design of the two test paper strips of single card, realizes through the sampling of same blood sample the utility model discloses detect influenza virus A type/B type and pneumonia chlamydia (english abbreviation is CP) and four kinds of pathogens of pneumonia mycoplasma (english abbreviation is MP) on the device simultaneously to realize once opening list, once sampling, detect four kinds of pathogens simultaneously, the clinical detection flow of simplification that can be very big, for patient's quick definite diagnosis, reasonable medicine use provides maximum facility, has important clinical use value.
The technical scheme of the utility model is that the device for synchronously detecting different types of influenza virus and pneumonia IgM antibodies comprises a double-channel card shell; the test strip FluA & B and the test strip CP & MP are arranged in the double-channel card shell in parallel; the first sample adding hole and the second sample adding hole are arranged on the card shell and are respectively and independently communicated with the test strip FluA & B and the test strip CP & MP; the test strip FluA & B comprises a first sample pad pretreated by an anti-erythrocyte antibody, a first blood filtering pad, a FluA & B tracer spraying glass fiber pad, a FluA & B antigen coated nitrocellulose membrane, first water absorption paper and a first plastic bottom plate; a first sample pad, a first blood filtering pad, a FluA & B tracer spraying glass fiber pad, a FluA & B antigen coated nitrocellulose membrane and first absorbent paper are sequentially adhered on the first plastic base plate; an influenza virus A detection line, an influenza virus B detection line and a first quality control line are sequentially arranged on the nitrocellulose membrane coated by the FluA & B antigens at intervals from the end of the FluA & B tracer spraying glass fiber pad to the end of the first absorbent paper, the influenza virus A detection line is coated with a high-specificity influenza virus A antigen, the influenza virus B detection line is coated with a high-specificity influenza virus B antigen, and the first quality control line is coated with a quality control line coated receptor; the first sample adding hole corresponds to the position of the first sample pad pretreated by the anti-erythrocyte antibody on the double-channel cartridge shell; the test strip CP & MP comprises a second sample pad pretreated by an anti-erythrocyte antibody, a second blood filtering pad, a CP & MP tracer spraying glass fiber pad, a CP & MP antigen coated nitrocellulose membrane, second absorbent paper and a second plastic base plate, wherein the second sample pad pretreated by the anti-erythrocyte antibody, the second blood filtering pad, the CP & MP tracer spraying glass fiber pad, the CP & MP antigen coated nitrocellulose membrane and the second absorbent paper are sequentially adhered to the second plastic base plate; a chlamydia pneumoniae detection line, a mycoplasma pneumoniae detection line and a second quality control line are sequentially arranged on the nitrocellulose membrane coated by the CP & MP antigen from the end of the CP & MP tracer spraying glass fiber pad to the end of the second water absorption paper at intervals, the chlamydia pneumoniae detection line is coated with a high-specificity chlamydia pneumoniae antigen, the mycoplasma pneumoniae detection line is coated with the high-specificity mycoplasma pneumoniae antigen, and the second quality control line is pre-coated with a quality control line coated receptor; the second sample adding hole corresponds to the position of the second sample pad pretreated by the anti-erythrocyte antibody on the double-channel cartridge shell.
Furthermore, two ends of the nitrocellulose membrane coated by the FluA & B antigen are respectively in staggered lap joint with the first absorbent paper and the FluA & B tracer spraying glass fiber pad, the other end of the FluA & B tracer spraying glass fiber pad is in staggered lap joint with the first blood filtering pad, and the other end of the first blood filtering pad is in staggered lap joint with the first sample pad; the two ends of the nitrocellulose membrane coated by the CP & MP antigen are respectively overlapped with the second absorbent paper and the CP & MP tracer spraying glass fiber pad in a staggered manner, the other end of the CP & MP tracer spraying glass fiber pad is overlapped with the second blood filtering pad in a staggered manner, and the other end of the second blood filtering pad is overlapped with the second sample pad in a staggered manner.
Furthermore, the FluA & B tracer spraying glass fiber mat and the CP & MP tracer spraying glass fiber mat are both prepared by diluting the marked tracer with a tracer buffer solution to obtain a tracer solution and then spraying glass fibers with the tracer solution; wherein the labeled tracer is obtained by labeling an anti-human IgM mu chain antibody and a quality control line labeled ligand with the tracer.
Furthermore, the tracer in the tracer-labeled anti-human IgM mu chain antibody and the quality control line-labeled ligand is one or a combination of more of colloidal gold, colloidal selenium, colored latex microspheres, fluorescent dye, fluorescent microspheres, quantum dots and carbon black particles; the particle size of the tracer particles is 30-300 nm.
Furthermore, the anti-human IgM mu chain antibody is a polyclonal antibody or a monoclonal antibody; when the anti-human IgM mu chain antibody is polyclonal, it includes sheep anti-human IgM mu chain, rabbit anti-human IgM mu chain, donkey anti-human IgM mu chain.
Furthermore, the tracer buffer solution comprises Tris-HCl solution, sucrose, trehalose and bovine serum albumin BSA, and has a pH value of 8.5, wherein the concentration of Tris-HCl is 0.02-0.1mol/L, the concentration of sucrose is 5-20% by mass, the concentration of trehalose is 1-5% by mass, and the concentration of bovine serum albumin BSA is 0.5-2% by mass.
Further, the first sample pad pretreated by the anti-erythrocyte antibody and the second sample pad pretreated by the anti-erythrocyte antibody are obtained by sequentially soaking and drying by using a pretreatment solution, wherein the treatment solution consists of Tris-HCL solution, the anti-erythrocyte antibody, bovine serum albumin BSA, casein and a surfactant Triton X-100, the concentration of the Tris-HCL solution is 0.1mol/L, the dilution degree of the anti-erythrocyte antibody is 1/200-1/2000, the concentration of the bovine serum albumin BSA is 0.5-1% by mass, the concentration of the casein is 0.1-0.2% by mass, and the concentration of the surfactant Triton X-100 is 0.5-1% by mass.
Still further, the first quality control line and the second quality control line are both coated with coating receptors, and the receptors coated on the first quality control line are matched with tracer substance marking line marking ligands in the FluA & B tracer spraying glass fiber mat; the receptor coated on the second quality control line is matched with a tracer marking quality control line marking ligand in the CP & MP tracer spraying glass fiber mat; the receptor-ligand pairing comprises rabbit/sheep anti-mouse IgG, rabbit anti-mouse IgG, sheep anti-chicken IgY, streptavidin-biotin coupling BSA and Protein A-mouse IgG.
Further, the first sample adding hole and the second sample adding hole are used for adding detection samples to the test strip FluA & B and the test strip CP & MP respectively, and the detection samples added to the first sample adding hole and the second sample adding hole are the same sample; the detection sample is whole blood, serum or plasma, and the whole blood is fingertip blood or venous blood; the volume of the detection sample required by each of the first sample adding hole and the second sample adding hole is 5-30 ul.
The utility model discloses an above-mentioned device of synchronous detection different grade type influenza virus and pneumonia IgM antibody carries out influenza virus A type, B type and pneumonia chlamydia, pneumonia mycoplasma's IgM antibody joint detection's method, including following step:
firstly, obtaining a detection sample according to a clinical sample collection standard;
secondly, simultaneously using matched sample diluent in a combined manner, namely adding the sample diluent to each sample adding hole while adding the detection sample to the first sample adding hole and the second sample adding hole S2, wherein the use amount of the sample diluent is 70-95 ul/sample adding hole;
thirdly, timing for 10-15 minutes after the sample adding is finished;
fourthly, immediately observing the line development conditions of the influenza A virus detection line, the influenza B virus detection line and the first quality control line on the test strip FluA & B after timing is finished; and the test paper strip CP & MP shows the line condition of the chlamydia pneumoniae detection line, the mycoplasma pneumoniae detection line and the second quality control line, and influenza virus A type, B type, chlamydia pneumoniae and mycoplasma infection results are obtained.
Wherein the sample diluent in the second step consists of Tris-HCL solution, casein, HAMA antibody inhibitor and surfactant Tween-20, wherein the concentration of the Tris-HCL solution is 0.1mol/L, the dilution degree of the HAMA antibody inhibitor is 1/1000-1/10000, the concentration of the casein is 0.2-2.0% by mass percent, and the concentration of the surfactant Tween-20 is 0.5-1% by mass percent.
The utility model discloses compare in prior art's advancement and lie in:
1. the utility model discloses limitation and pain point to clinical virus detects and pneumonia chlamydia mycoplasma detects, a calorie of two allies oneself with four indexs joint detection device has been designed, this joint detection device includes test paper strip FluA & B that influenza A virus type/B type detected, and the test paper strip CP & MP that pneumonia chlamydia/mycoplasma detected, two test paper strips set up respectively in the binary channels of joint device's binary channels card shell, mutual isolation in a card shell, when realizing the synchronous independent detection of four indexs, the sample of two test paper strips, the testing process, the testing result is mutually independent, each other does not influence, finally realized that influenza A virus type/B type and pneumonia chlamydia/mycoplasma's detection is unified simply on a card, fast, go on in step.
2. The utility model discloses envelope antigen on the detection line on the nitrocellulose membrane among the composite set, the envelope antibody on the matter accuse line realizes the IgM antibody detection: an influenza virus A detection line, an influenza virus B detection line and a first quality control line are arranged on the first antigen-coated nitrocellulose membrane at intervals, the influenza virus A detection line is coated with a high-specificity influenza virus A antigen, the influenza virus B detection line is coated with a high-specificity influenza virus B antigen, and the first quality control line is coated with a quality control line-coated receptor; and a chlamydia pneumoniae detection line, a mycoplasma pneumoniae detection line and a second quality control line are arranged on the nitrocellulose membrane coated with the second antigen at intervals, the chlamydia pneumoniae detection line is coated with a high-specificity chlamydia pneumoniae antigen, the mycoplasma pneumoniae detection line is coated with a high-specificity mycoplasma pneumoniae antigen, and the second quality control line is coated with a solid phase quality control line coated receptor. Different from the antigen detection and other detection indexes in the prior art, the kit has more clinical judgment accuracy.
3. Based on the combined detection device of the utility model, the detection method provided by the utility model can synchronously obtain the detection results of influenza virus A/B and Chlamydia Pneumoniae (CP)/mycoplasma through single sample sampling and synchronous assay, only adopt a blood sample to detect four kinds of pathogens promptly, compare in prior art and need take the nasopharynx swab as sample detection in order to obtain influenza virus A type/B type, take blood as sample detection appearance and obtain the testing result of Chlamydia Pneumoniae (CP)/mycoplasma, the utility model discloses a once open the list, once sample, detect four kinds of pathogens simultaneously, practiced thrift detection cost, patient financial resources and time, improved diagnostic efficiency, greatly simplify the clinical detection flow, for patient's quick definite diagnosis, rationally use medicine provides the biggest facility, has important clinical use value.
4. The utility model discloses the device can detect serum, and plasma can detect whole blood again, can detect venous blood, and the preferred fingertip blood that can detect, and it is few to detect required blood sample quantity, and this provides very big facility to clinical application, and the reason has two: firstly, as is well known, the old and children are susceptible and high-incidence people with respiratory diseases, but the old is aged due to blood vessels, and the children, especially infants are extremely thin due to blood vessels, so that the collection of venous blood of the two people is difficult and troublesome in clinical practice, the finger tip blood can be detected, and the sampling simplification of the two people has great significance; it is two, respiratory disease is especially the utility model discloses the respiratory disease that these four kinds of pathogens that detection device aimed at arouse, the epidemic phase morbidity is concentrated, and the symptom is often heavier, unusual misery of feeling when the patient is hospitalized, and easy cross infection, traditional serum, plasma detection is because need carry out centrifugal treatment to the blood sample after gathering, the detection time has increased in the intangible, and then the time that the patient waited has been increased, the process of waiting when increasing the misery has also increased cross infection's risk, and fingertip blood detects, it is simple and convenient quick, need not unnecessary sample processing, prick one drop, the result has just come out within 10-15 minutes, this has very big clinical application and worth undoubtedly.
Drawings
These and/or other aspects and advantages of the present invention will become more apparent and more readily appreciated from the following detailed description of the embodiments of the invention, taken in conjunction with the accompanying drawings of which:
FIG. 1 is a graph showing the trend of IgM and IgG antibodies appearing after infection with a pathogen in the background of the present invention;
FIG. 2 is a schematic view of the overall structure of an apparatus for synchronously detecting different types of influenza viruses and pneumonia IgM antibodies in an embodiment of the present invention;
fig. 3 is a schematic structural composition diagram of a test strip FluA & B and a test strip CP & MP of the device for synchronously detecting different types of influenza viruses and pneumonia IgM antibodies in the embodiment of the present invention;
FIG. 4 is a schematic diagram of different detection results of influenza A and B viruses obtained by the device for synchronously detecting different types of influenza viruses and pneumonia IgM antibodies in the embodiment of the present invention;
fig. 5 is a schematic diagram of different detection results of chlamydia pneumoniae and mycoplasma obtained by the device for synchronously detecting different types of influenza viruses and pneumonia IgM antibodies in the embodiment of the present invention.
Detailed Description
In order to better understand the present invention for those skilled in the art, the present invention will be described in further detail with reference to the accompanying drawings and specific embodiments.
Example 1
A combined device for synchronously detecting IgM antibodies of influenza virus A type, B type, Chlamydia pneumoniae and Mycoplasma has a structure shown in FIG. 2 and comprises a double-channel card shell; the test strip FluA & B and the test strip CP & MP are arranged in the double-channel card shell in parallel; and the first sample adding hole S1 and the second sample adding hole S2 are arranged on the card shell and are respectively and independently communicated with the test strip FluA & B and the test strip CP & MP. The specific design of each part is as follows:
the structure of the test strip FluA & B is shown in figure 3, and comprises a first sample pad 1 pretreated by an anti-erythrocyte antibody, a first blood filtering pad 2, a FluA & B tracer spraying glass fiber pad 3, a FluA & B antigen coated nitrocellulose membrane 4, a first absorbent paper 5 and a first plastic bottom plate 6; paste first sample pad 1, first filter blood pad 2, FluA & B tracer spraying glass fiber pad 3, FluA & B antigen coated nitrocellulose membrane 4, first absorbent paper 5 on the first plastic bottom plate 6 in proper order, and each part connects the preferred design and is: the two ends of a nitrocellulose membrane 4 coated by the FluA & B antigen are respectively in staggered lap joint with a first absorbent paper 5 and a FluA & B tracer spraying glass fiber 3, the other end of the FluA & B tracer spraying glass fiber pad 3 is in staggered lap joint with a first blood filtering pad 2, and the other end of the first blood filtering pad 2 is in staggered lap joint with a first sample pad 1 pretreated by an anti-erythrocyte antibody; an influenza virus A detection line A, an influenza virus B detection line B and a first quality control line C are sequentially arranged on the nitrocellulose membrane 4 coated with the FluA & B antigens at intervals from the end 3 of the FluA & B tracer spraying glass fiber pad to the end 5 of the first absorbent paper, the influenza virus A detection line A is coated with the high-specificity influenza virus A antigens, the influenza virus B detection line B is coated with the high-specificity influenza virus B antigens, and the first quality control line C is coated with the quality control line coated receptors; the first sample application hole S1 corresponds to the position of the first sample pad 1 pretreated with the anti-erythrocyte antibody on the two-channel cartridge.
The test strip CP & MP is structurally shown in fig. 3, and includes a second sample pad 1 'pretreated by anti-erythrocyte antibody, a second blood filter pad 2', a CP & MP tracer spray glass fiber pad 3 ', a CP & MP antigen coated nitrocellulose membrane 4', a second absorbent paper 5 'and a second plastic base plate 6', the second sample pad 1 'pretreated by anti-erythrocyte antibody, the second blood filter pad 2', the CP & MP tracer spray glass fiber pad 3 ', the CP & MP antigen coated nitrocellulose membrane 4', and the second absorbent paper 5 'are sequentially adhered to the second plastic base plate 6', and the connection of each part is preferably designed as: the two ends of a nitrocellulose membrane 4 ' coated by a CP & MP antigen are respectively in staggered lap joint with a second absorbent paper 5 ' and a CP & MP tracer spraying glass fiber pad 3 ', the other end of the CP & MP tracer spraying glass fiber pad 3 ' is in staggered lap joint with a second blood filtering pad 2 ', and the other end of the second blood filtering pad 2 ' is in staggered lap joint with a second sample pad 1 ' pretreated by an anti-erythrocyte antibody; a chlamydia pneumoniae detection line CP, a mycoplasma pneumoniae detection line MP and a second quality control line C ' are sequentially arranged on the nitrocellulose membrane 4 ' coated with the CP & MP antigens at intervals from the 3 ' end of the CP & MP tracer spraying glass fiber pad to the 5 ' end of the second absorbent paper, the chlamydia pneumoniae detection line CP is coated with high-specificity chlamydia pneumoniae antigens, the mycoplasma pneumoniae detection line MP is coated with high-specificity mycoplasma pneumoniae antigens, and the second quality control line C ' is pre-coated with a quality control line coated receptor; the second sample application hole S2 corresponds to the position of the second sample pad 1' pretreated with the anti-erythrocyte antibody on the two-channel cartridge.
Wherein: the FluA & B tracer spraying glass fiber mat 3 and the CP & MP tracer spraying glass fiber mat 3' are both prepared by diluting a labeled tracer with a tracer buffer solution to obtain a tracer solution and then spraying glass fibers with the tracer solution; wherein the labeled tracer is obtained by labeling an anti-human IgM mu chain antibody and a quality control line labeled ligand with the tracer. Preferably, the tracer in the tracer marked anti-human IgM mu chain antibody and the quality control line marked ligand is one or a combination of more of colloidal gold, colloidal selenium, colored latex microspheres, fluorescent dye, fluorescent microspheres, quantum dots and carbon black particles; the particle size of the tracer particles is 30-300 nm. Also included are preferred designs in which: the anti-human IgM mu chain antibody is a polyclonal antibody or a monoclonal antibody; when the anti-human IgM mu chain antibody is a polyclonal antibody, it includes sheep anti-human IgM mu chain, rabbit anti-human IgM mu chain, donkey anti-human IgM mu chain; the tracer buffer solution consists of Tris-HCl solution, sucrose, trehalose and bovine serum albumin BSA, and has a pH value of 8.5, wherein the concentration of Tris-HCl is 0.02-0.1mol/L, the concentration of sucrose is 5-20% by mass, the concentration of trehalose is 1-5% by mass, and the concentration of bovine serum albumin BSA is 0.5-2% by mass.
In the embodiment, the first sample pad 1 pretreated by the anti-erythrocyte antibody and the second sample pad 1' pretreated by the anti-erythrocyte antibody are obtained by sequentially soaking and drying by using a pretreatment solution, wherein the treatment solution consists of Tris-HCL solution, the anti-erythrocyte antibody, bovine serum albumin BSA, casein and a surfactant Triton X-100, the concentration of the Tris-HCL solution is 0.1mol/L, the dilution degree of the anti-erythrocyte antibody is 1/200-1/2000, the concentration of the bovine serum albumin BSA is 0.5-1% by mass, the concentration of the casein is 0.1-0.2% by mass, and the concentration of the surfactant Triton X-100 is 0.5-1% by mass.
In the aspect of designing a quality control line and a tracer marking quality control line marking ligand: coating receptors are coated on the first quality control line C and the second quality control line C', and the receptors coated on the first quality control line C are matched with tracer substance marking quality control line marking ligands in the FluA & B tracer spraying glass fiber mat 3; the receptor coated on the second quality control line C 'is matched with a tracer marking quality control line marking ligand in the CP & MP tracer spraying glass fiber mat 3'; the pairing comprises rabbit/sheep anti-mouse IgG, rabbit anti-mouse IgG, sheep anti-chicken IgY, streptavidin-biotin coupling BSA and Protein A-mouse IgG.
In the combination device of this embodiment, the first well S1 and the second well S2 are used for adding the detection samples to the test strip FluA & B and the test strip CP & MP, respectively, and the detection samples added to the first well S1 and the second well S2 are the same sample; the detection sample is whole blood, serum or plasma, and the whole blood is fingertip blood or venous blood; the volume of the detection sample required by each of the first sample application well S1 and the second sample application well S2 is 5-30 ul.
Example 2
The method for preparing the combined device for synchronously detecting the IgM antibodies of the influenza viruses of the A type, the B type, the Chlamydia pneumoniae and the Mycoplasma comprises the following steps:
a) preparing a tracer: tracers include, but are not limited to, colloidal gold, colloidal selenium, colored latex microspheres, fluorescent dyes, fluorescent microspheres, quantum dots, carbon black particles, and the like;
b) marking an anti-human IgM mu chain antibody and a quality control line marking ligand by adopting the tracer prepared in the step a) to obtain a marked tracer;
c) diluting the labeled tracer in the step b) by adopting a tracer buffer solution to obtain a tracer solution, and spraying the tracer solution on glass fiber to prepare a tracer-sprayed glass fiber mat;
d) respectively spraying a piece of nitrocellulose membrane with a pre-diluted high-specificity influenza virus A, B as an influenza virus A detection line and an influenza virus B detection line, respectively spraying a point quality control line coated antibody as a quality control line, respectively spraying a chlamydia pneumoniae antigen and a mycoplasma pneumoniae antigen as a chlamydia pneumoniae detection line CP and a mycoplasma pneumoniae detection line MP on the other nitrocellulose membrane, respectively preparing a FluA & B antigen coated nitrocellulose membrane and a CP & MP antigen coated nitrocellulose membrane by using a point quality control line coated receptor as quality control lines;
e) the first sample pad 1 pretreated with anti-erythrocyte antibody and the second sample pad 1' pretreated with anti-erythrocyte antibody are processed: soaking and drying by using a pretreatment solution, wherein the treatment solution consists of Tris-HCL solution, anti-erythrocyte antibody, bovine serum albumin BSA, casein and surfactant Triton X-100, the concentration of the Tris-HCL solution is 0.1mol/L, the dilution degree of the anti-erythrocyte antibody is 1/200-1/2000, the concentration of the bovine serum albumin BSA is 0.5-1% by mass percent, the concentration of the casein is 0.1-0.2% by mass percent, and the concentration of the surfactant Triton X-100 is 0.5-1% by mass percent;
f) sequentially and respectively sticking the pretreated sample pad obtained in the step e), the untreated blood filter pad, the tracer spray glass fiber pad prepared in the step c), the cellulose nitrate membrane prepared in the step d) and the absorbent paper on respective plastic base plates to respectively prepare test paper strips FluA & B and CP & MP, and finally respectively putting the two test paper strips into a two-connected plastic card shell.
Theoretically: because the two test strips are independent, the receptors coated by the quality control lines on the two test strips can be matched with the tracer substance mark quality control line mark ligands in the corresponding tracer substance spraying glass fiber mat in the same or different modes, and no problem exists at all. Therefore, the tracer labeled anti-human IgM mu chain antibody and the quality control line labeled ligand used in spraying the glass fiber mat with the tracer on the test strip FluA & B and the test strip CP & MP in the step B) above may be the same or different, as long as the tracer labeled quality control line labeled ligand used is paired with the receptor coated with the quality control line on the nitrocellulose membrane on the same test strip. Similarly, the proportions of the components in the treatment solution formulas of the first sample pad pretreated by the anti-erythrocyte antibody on the test strip FluA & B and the second sample pad pretreated by the anti-erythrocyte antibody on the test strip CP & MP can be the same or different.
Example 3
Detection method and result interpretation
A method for simultaneous detection of IgM antibodies of influenza virus type a, type B and chlamydia pneumoniae, mycoplasma pneumoniae using the combination of IgM antibodies of influenza virus type a, type B and chlamydia pneumoniae, mycoplasma pneumoniae of example 1, comprising the steps of:
firstly, obtaining a detection sample according to a clinical sample collection standard;
secondly, simultaneously using matched sample diluents in a combined manner, namely adding the sample diluents to each sample adding hole while adding the detection samples to the first sample adding hole S1 and the second sample adding hole S2, wherein the usage amount of the sample diluents is 70-95 ul/sample adding hole;
thirdly, timing for 10-15 minutes after the sample adding is finished;
fourthly, immediately observing the color change conditions of an influenza virus A detection line A, an influenza virus B detection line B and a first quality control line C on the test strip FluA & B after timing is finished; and the color change conditions of the chlamydia pneumoniae detection line CP, the mycoplasma pneumoniae detection line MP and the second quality control line C' on the test paper strip CP & MP are obtained, so that infection results of influenza viruses A, B, chlamydia pneumoniae and mycoplasma are obtained.
Wherein the sample diluent in the second step consists of Tris-HCL solution, casein, HAMA antibody inhibitor and surfactant Tween-20, wherein the concentration of the Tris-HCL solution is 0.1mol/L, the dilution degree of the HAMA antibody inhibitor is 1/1000-1/10000, the concentration of the casein is 0.2-2.0% by mass percent, and the concentration of the surfactant Tween-20 is 0.5-1% by mass percent.
And (4) interpretation of results:
for the test strip FluA & B, the different line-showing results in FIG. 4, which may appear in 8, are interpreted as follows:
1. if the first quality control line C shows line and the influenza A detection line A and/or the influenza B detection line B simultaneously shows line, three conditions are distinguished:
1) the first quality control line C shows line + influenza A virus detection line A shows line + influenza B virus detection line B shows line, which indicates mixed infection influenza A virus and B virus;
2) the first quality control line C shows line and the influenza A virus detection line A shows line, which indicates that the influenza A virus is infected;
3) the first quality control line C shows a line + + influenza B virus detection line B shows that influenza B virus is infected;
2. if only the first quality control line C shows line, other detection lines do not show line, which indicates that the detection results of the influenza virus infection type A, B are all negative;
3. if the first quality control line C is not displayed or no line is displayed on the test strip, the detection result is invalid, and the retest is required.
For the test strip CP & MP, the possible line-development results are shown in fig. 5, which is basically consistent with the detection results and interpretation methods of the test strip FluA & B.
It can be seen that the utility model discloses joint detection device and detection method with the gathering of influenza virus A, B type and four kinds of pathogens of pneumonia chlamydia, mycoplasma pneumoniae on a detection card, realized once gathering blood sample, realize the IgM antibody detection that four kinds of pathogens acute phase infects the appearance simultaneously.
While various embodiments of the present invention have been described above, the above description is intended to be illustrative, not exhaustive, and not limited to the disclosed embodiments. Many modifications and variations will be apparent to those of ordinary skill in the art without departing from the scope and spirit of the described embodiments. Therefore, the protection scope of the present invention shall be subject to the protection scope of the claims.
Claims (7)
1. A device for synchronously detecting different types of influenza viruses and pneumonia IgM antibodies is characterized by comprising a double-channel card shell; the test strip FluA & B and the test strip CP & MP are arranged in the double-channel card shell in parallel; a first sample adding hole (S1) and a second sample adding hole (S2) which are respectively and independently communicated with the test strip FluA & B and the test strip CP & MP on the card shell;
the test strip FluA & B comprises a first sample pad (1) pretreated by an anti-erythrocyte antibody, a first blood filter pad (2), a FluA & B tracer spraying glass fiber pad (3), a FluA & B antigen coated nitrocellulose membrane (4), first absorbent paper (5) and a first plastic bottom plate (6); a first sample pad (1), a first blood filtering pad (2), a FluA & B tracer spraying glass fiber pad (3), a FluA & B antigen coated nitrocellulose membrane (4) and first absorbent paper (5) are sequentially adhered to the first plastic bottom plate (6); an influenza virus A detection line (A), an influenza virus B detection line (B) and a first quality control line (C) are sequentially arranged on the nitrocellulose membrane (4) coated by the FluA & B antigens at intervals from the end of the FluA & B tracer spraying glass fiber pad (3) to the end of the first absorbent paper (5), the influenza virus A detection line (A) is coated with a high-specificity influenza virus A antigen, the influenza virus B detection line (B) is coated with a high-specificity influenza virus B antigen, and the first quality control line (C) is coated with a quality control line coated receptor; the first application well (S1) corresponds to a position of the first sample pad (1) pretreated with the anti-erythrocyte antibody on the two-channel cartridge;
the test strip CP & MP comprises a second sample pad (1 ') pretreated by an anti erythrocyte antibody, a second blood filtering pad (2'), a CP & MP tracer spraying glass fiber pad (3 '), a nitrocellulose membrane (4') coated by a CP & MP antigen, second absorbent paper (5 ') and a second plastic bottom plate (6'), wherein the second sample pad (1 ') pretreated by the anti erythrocyte antibody, the second blood filtering pad (2'), the CP & MP tracer spraying glass fiber pad (3 '), the nitrocellulose membrane (4') coated by the CP & MP antigen and the second absorbent paper (5 ') are sequentially stuck on the second plastic bottom plate (6'); a chlamydia pneumoniae detection line (CP), a mycoplasma pneumoniae detection line (MP) and a second quality control line (C ') are sequentially arranged on the nitrocellulose membrane (4 ') coated by the CP & MP antigen from the end of the CP & MP tracer spraying glass fiber pad (3 ') to the end of the second absorbent paper (5 ') at intervals, the chlamydia pneumoniae detection line (CP) is coated with a high-specificity chlamydia pneumoniae antigen, the mycoplasma pneumoniae detection line (MP) is coated with the high-specificity mycoplasma pneumoniae antigen, and the second quality control line (C ') is pre-coated with a quality control line coated receptor; the second application hole (S2) corresponds to the position of the second sample pad (1') pretreated with the anti-erythrocyte antibody on the two-channel cartridge.
2. The apparatus for simultaneous detection of different types of influenza virus and pneumonia IgM antibodies according to claim 1,
the two ends of the nitrocellulose membrane (4) coated by the FluA & B antigens are respectively in staggered lap joint with the first absorbent paper (5) and the FluA & B tracer spraying glass fiber pad (3), the other end of the FluA & B tracer spraying glass fiber pad (3) is in staggered lap joint with the first blood filtering pad (2), and the other end of the first blood filtering pad (2) is in staggered lap joint with the first sample pad (1) pretreated by the anti-erythrocyte antibodies.
3. The apparatus for simultaneous detection of different types of influenza virus and pneumonia IgM antibodies according to claim 2,
the two ends of the nitrocellulose membrane (4 ') coated by the CP & MP antigen are respectively in staggered lap joint with the second absorbent paper (5 ') and the CP & MP tracer spraying glass fiber pad (3 '), the other end of the CP & MP tracer spraying glass fiber pad (3 ') is in staggered lap joint with the second blood filtering pad (2 '), and the other end of the second blood filtering pad (2 ') is in staggered lap joint with the second sample pad (1 ') pretreated by the anti-erythrocyte antibody.
4. The device for the simultaneous detection of IgM antibodies of different types of influenza viruses and pneumonia according to claim 3, wherein said FluA & B tracer-coated glass fiber mat (3) and CP & MP tracer-coated glass fiber mat (3') each comprise a tracer and its labeled anti-human IgM μ chain antibody and quality control line labeled ligand; the particle size of the tracer particles is 30-300 nm.
5. The device for synchronously detecting the IgM antibodies of different types of influenza viruses and pneumonia according to claim 4, wherein said anti-human IgM mu chain antibody is a polyclonal antibody or a monoclonal antibody; when the anti-human IgM mu chain antibody is a polyclonal antibody, it includes one of sheep anti-human IgM mu chain, rabbit anti-human IgM mu chain and donkey anti-human IgM mu chain.
6. The apparatus for simultaneous detection of different types of influenza virus and pneumonia IgM antibodies according to claim 5,
the first quality control line (C) and the second quality control line (C') are both coated with coating receptors, and the coating receptors on the first quality control line (C) are matched with tracer marking quality control line marking ligands in the FluA & B tracer spraying glass fiber mat (3); the coated receptors on the second control line (C ') are paired with tracer-labeled control line-labeled ligands in the CP & MP tracer-sprayed glass fiber mat (3'); the coating receptor and the tracer marker quality control line marker ligand are matched, including the matching of rabbit/sheep anti-mouse IgG and mouse IgG, the matching of rabbit anti-mouse IgG and mouse IgG, the matching of sheep anti-chicken IgY and chicken IgY, the matching of streptavidin and biotin coupling BSA, and the matching of Protein A and mouse IgG.
7. The apparatus for synchronously detecting IgM antibodies of different types of influenza viruses and pneumonia according to claim 1, wherein the first well (S1) and the second well (S2) are used for adding a detection sample to a test strip FluA & B and a test strip CP & MP, respectively, and the detection samples added to the first well (S1) and the second well (S2) are homogeneous samples; the detection sample is whole blood, serum or plasma, and the whole blood is fingertip blood or venous blood; each of the first well (S1) and the second well (S2) requires a volume of the test sample of 5 to 30 ul.
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