CN219957599U - Quick six-joint detection immunochromatography test strip for carbapenemase - Google Patents

Abstract

The utility model discloses a carbapenemase rapid six-joint detection immunochromatography test strip which mainly comprises a bottom plate, a partition plate, an NC film, a sample pad, a gold mark/milk mark pad and an absorption pad; the NC film is fixed on the bottom plate, the gold mark/milk mark pad and the absorption pad are respectively fixed at two ends of the NC film, the sample pad is lapped on the gold mark/milk mark pad, the partition plate is positioned in the middle of the NC film and is perpendicular to the NC film for fixation, and the left end of the partition plate is positioned at the joint of the sample pad and the gold mark/milk mark pad; three detection lines and a quality control line are respectively arranged on NC films at two sides of the partition board, monoclonal antibodies of the carbapenemases are respectively coated on the six detection lines, goat anti-mouse IgG antibodies are coated on the quality control line, and 3 kinds of monoclonal antibodies marked by colloidal gold/latex microspheres are respectively sprayed on gold/latex pads at two sides. The test strip can detect gram-negative bacillus with carbapenems resistance common clinically at the same time, has high sensitivity and specificity, and can obtain a visual result within 15 min.

Description

Quick six-joint detection immunochromatography test strip for carbapenemase

Technical Field

The utility model relates to the field of biological detection, in particular to a carbapenemase rapid six-joint detection immunochromatography test strip.

Background

The carbapenem drug resistance rate of clinical pathogenic bacteria is high, and the preparation method brings great challenges to the global medical and health industries and the life quality of human beings. How to quickly and accurately identify the drug-resistant bacteria becomes a difficult problem of clinical treatment at the present stage, and has important influences on the hospitalization time, prognosis, medical cost and the like of patients. The drug-resistant mechanism of carbapenem-resistant gram-negative bacteria is mainly beta-lactamase, wherein five carbapenemases are most common, and the five carbapenemases are respectively: KPC (Klebsiella pneumoniae carbapenemases), NDM (new delhi metallo-. Beta. -lactase), VIM (verona integer-encoded metal lo-. Beta. -lactase), IMP (imipenemase metallo-. Beta. -lactase) and OXA (oxacillinase carbapenemases). Wherein OXA-48 is commonly found in bacteria of the order Enterobacteriaceae and OXA-23 is commonly found in Acinetobacter baumannii. The corresponding therapeutic drugs of different types of enzymes are different, if the six enzymes can be identified as early as possible, most carbapenem-resistant gram-negative bacteria can be identified, and meanwhile, the proper antibiotics can be determined, so that a clinician can make decisions as early as possible, the drugs are reasonably used, first-line vitality is strived for patients, and the spread of drug-resistant bacteria is prevented in time.

Immunochromatography (immunochromatography assay, ICA) has become a popular detection method in the biomedical detection field, and has the advantages of simple operation, quick response, visual result, low requirements on equipment environment, and good application and development in the bedside detection field. However, under the limitation of accuracy and sensitivity, the detection of the carbapenemase type is still limited to 2-5 proteins at the present stage, so that the structure of the detection carrier is designed and optimized on the basis of the prior art, and the detection efficiency, accuracy and sensitivity of the six-joint detection test strip can reach good levels on detecting six enzyme types KPC, NDM, VIM, IMP, OXA-23 and OXA-48.

Disclosure of Invention

Aiming at the defects of the current clinical carbapenem-resistant bacteria detection method, the utility model provides a carbapenemase rapid six-joint detection immunochromatography test strip, which realizes qualitative and semi-quantitative rapid synchronous detection of clinical common carbapenem-resistant gram-negative bacteria by using an immunochromatography technology, and can realize synchronous rapid detection of KPC, NDM, VIM, IMP, OXA-23 and OXA-48 six carbapenemases; the test strip is simple to operate, convenient to carry and short in result interpretation time, and can be used under various conditions.

The technical scheme adopted for solving the technical problems is as follows:

the utility model firstly protects a carbapenemase rapid six-joint detection immunochromatography test strip, which mainly comprises a bottom plate, a partition plate, an NC film, a sample pad, a gold mark/milk mark pad and an absorption pad; the NC film is fixed on the bottom plate, the gold mark/milk mark pad and the absorption pad are respectively fixed at two ends of the NC film, the sample pad is lapped on the gold mark/milk mark pad, the partition plate is positioned in the middle of the NC film and is perpendicular to the NC film for fixation, and the left end of the partition plate is positioned at the joint of the sample pad and the gold mark/milk mark pad; three detection lines and a quality control line are respectively arranged on NC films at two sides of the partition board, monoclonal antibodies of the carbapenemases are respectively coated on the six detection lines, goat anti-mouse IgG antibodies are coated on the quality control line, and 3 kinds of monoclonal antibodies marked by colloidal gold/latex microspheres are respectively sprayed on gold/latex pads at two sides.

As a further preferred aspect of the present utility model, the monoclonal antibodies are KPC protein monoclonal antibody, NDM protein monoclonal antibody, VIM protein monoclonal antibody, IMP protein monoclonal antibody, OXA-23 protein monoclonal antibody and OXA-48 protein monoclonal antibody, respectively.

As a further preferred aspect of the present utility model, the NC films are two, the two NC films are fixed side by side on the base plate, and the separator is positioned between the two NC films and fixed perpendicular to the NC films. The two films can be respectively paved with 4 different antibodies, so that the technical difficulty of paving 7 antibodies by one film is reduced, and meanwhile, the interference effect between samples and between antibodies is avoided.

More preferably, each NC film is independently provided with a gold/milk label pad and an absorbent pad.

As a further preferable mode of the utility model, the test strip is sleeved with the plastic clamping shell, and is provided with a sample adding hole, a result observation window and ventilation holes.

More preferably, the number of the result observation windows and the ventilation holes is 2, and the result observation windows and the ventilation holes are independently positioned on two sides of the partition board. The width of the result observation window is set according to practical conditions, such as 4mm, 5mm, 6mm, 7mm, 8mm and the like

Further preferably, the detection lines and the quality control lines on the NC film are arranged at equal intervals, for example, the width and the interval may be set according to actual needs, for example, the width is set to 5mm, and the interval is set to 5mm.

As a further preferred aspect of the present utility model, the colloidal gold/latex microsphere has a particle size of 20 to 40nm, such as 20nm,25nm,30nm,35nm,40nm, etc.

Preferably, the test strip can be further filled into an aluminum foil packaging bag to form a test strip product.

In use, the sample to be tested may be a single isolated colony or a positive blood sample.

As a further preference of the utility model, the bottom plate is a PVC bottom plate, the partition plate is a PVC partition plate, the NC membrane is Millipore135 in U.S., the sample pad is a Ahlstrom 8964 glass cellulose membrane in Finland, the gold mark pad is a Ahlstrom 6613 polyester fiber membrane in Finland, and the absorption pad is cellulose filter paper.

Advantageous effects

The quick six-joint detection immunochromatography test strip for carbapenemases can realize synchronous and quick detection of KPC, NDM, VIM, IMP, OXA-23 and OXA-48 carbapenemases on one test strip, is simple to operate, does not need large-scale equipment and instruments, has low professional requirements on surrounding environment and detection personnel, has high detection speed, can obtain a visual result within 15 minutes, can provide information such as gram-negative bacillus carbapenemases for clinicians and researchers in time, greatly makes up the defect of clinical examination work at the present stage, and guides clinical work to timely and pertinently use antibiotics.

Drawings

FIG. 1 is a schematic diagram of immunochromatographic test strip detection;

FIG. 2 is a schematic diagram of the structural hierarchy of the novel immunochromatographic test strip;

FIG. 3 is a schematic diagram of a test strip;

FIG. 4 is a diagram of result interpretation;

1-sample pad; 2-gold/milk label pad; 3-PVC bottom plate; 4-NC film; 5-an absorbent pad; 6-a quality control line; 7-detecting lines; 71-detection line 1; 72-detecting line 2; 73-detection line 3; 74-detecting line 4; 75-detecting line 5; 76-detecting line 6; 8-a sample adding hole; 9-a result observation window; 10-ventilation holes; 11-PVC separator.

Detailed Description

In order that the present utility model may be more readily understood, a more particular description of the utility model will be rendered by reference to specific embodiments that are illustrated in the appended drawings. The utility model will now be described in further detail with reference to the accompanying drawings. The drawings are simplified schematic representations which merely illustrate the basic structure of the utility model and therefore show only the structures which are relevant to the utility model.

Example 1

Referring to fig. 2 and 3, a carbapenemase rapid six-joint detection immunochromatography test strip comprises a PVC bottom plate 3, NC films 4, a sample pad 1, a gold mark/milk mark pad 2, and an absorption pad 5, wherein the NC films 4 are fixed on the PVC bottom plate 3 in two pieces side by side, the PVC separator 11 is fixed between the two NC films and perpendicular to the NC films 4, the gold mark/milk mark pad 2 and the absorption pad 5 are respectively fixed at two ends of the NC films 4, and the sample pad 1 is lapped on the gold mark/milk mark pad 2; three detection lines 7 and a quality control line 6 are respectively arranged on the two NC films 4, monoclonal antibodies of the carbapenemases are respectively coated on the six detection lines 7, goat anti-mouse IgG antibodies are coated on the quality control line 6, and 3 kinds of monoclonal antibodies marked by colloidal gold/latex microspheres are respectively sprayed on gold marks/latex marks on two sides;

wherein, the detection line 71, the detection line 72, the detection line 73 and one of the quality control lines on the NC film 4 are arranged at equal intervals, the width is 5mm, and the interval is 5mm; the detection lines 74, 75 and 76 are arranged at equal intervals with another quality control line.

Wherein, the monoclonal antibodies of the carbapenemases are KPC protein monoclonal antibody, NDM protein monoclonal antibody, VIM protein monoclonal antibody, IMP protein monoclonal antibody, OXA-23 protein monoclonal antibody and OXA-48 protein monoclonal antibody respectively.

In further embodiments, the colloidal gold/latex microsphere marking particle size is 20 to 40nm.

In other embodiments, the test strip is sleeved with a plastic clamping shell and is provided with a sample adding hole 8, a result observation window 9 and an air hole 10; more specifically, the number of the result observation windows 9 and the number of the ventilation holes 10 are 2, and the result observation windows and the ventilation holes 10 are independently located at two sides of the partition board, and the shape of the observation windows is not limited, for example, the observation windows can be rectangular, and the width can be set according to practical conditions, for example, the observation windows can be 5mm.

In further embodiments, the sample to be tested may be an isolated single colony or a positive blood sample.

In other embodiments, the substrate is a PVC substrate with an adhesive, the NC film is Millipore135, the sample pad is a glass cellulose film of Ahlstrom 8964, finland, the gold mark pad is a polyester fiber film of Ahlstrom 6613, finland, and the absorbent pad is cellulose filter paper.

Example 2

Based on the embodiment 1, the utility model prepares a carbapenemase rapid six-joint detection immunochromatography test strip. The preparation process comprises the following steps:

(1) The preparation method of the immunochromatography test strip for rapid six-up detection of carbapenemase comprises the following steps:

(A) NC membrane antibody coating

Coating six commercial monoclonal antibodies on six detection lines of an NC film by using a BIO DOT spot film instrument, coating a goat anti-mouse monoclonal antibody on a quality control line, and closing the rest blank sites;

(B) Preparation of gold/milk label pad

Putting the polyester fiber film into 10% BSA blocking solution, soaking for a period of time, taking out, drying in a 37 ℃ oven for 5 hours, spraying the prepared six gold-labeled/milk-labeled antibodies on the dried polyester fiber film by using a BIO DOT spot film tester, and preserving at 4 ℃ for later use;

(C) Preparation of sample pad

Soaking a glass cellulose membrane in a 10% BSA blocking solution, taking out, drying in a 37 ℃ oven for 5 hours, and preserving in a room temperature drying environment for later use;

(D) Assembly of test strips

The first layer is a PVC bottom plate 3, the second layer is an NC film 4 which is divided into two pieces and separated by a PVC partition plate 11, three detection line antibodies (T1, T2, T3/T4, T5, T6) and a quality control antibody (C) are respectively coated on the two NC films, the third layer is a gold mark/milk mark pad 2, the fourth layer is a sample pad 1, the sample pad is used as a starting end, an absorption pad 5 is stuck at a terminal end, and the components are connected end to end and are sequentially stacked and assembled. The strips were cut into individual strips 5mm wide by a slitter and placed into special plastic card shells provided with sample application holes 8, result observation windows 9 and ventilation holes 10 as shown in fig. 3. Sealing, drying and preserving.

(2) Principle of detection

And (3) adding the sample into a sample adding hole, dissolving colloidal gold/latex microsphere labeled antibodies on a gold label/latex label pad by a sample solution, and forming a complex by combining the antigen with the gold label/latex label monoclonal antibodies specifically if the sample contains the antigen to be detected, wherein the complex is trapped by forming a gold label antibody-antigen-capture antibody sandwich complex with the solid-phase monoclonal antibodies at a detection line due to the upward chromatography effect, and the colloidal gold/latex microsphere is gathered in a large amount to be red, so that the color depth is related to the amount of the antigen to be detected. And the excessive gold-labeled antibody continues to ascend, and is combined with goat anti-mouse IgG at a quality control line to aggregate, precipitate and develop color, so that the chromatography is smooth, and the detection result is effective.

(3) Specific detection method

(A) Treatment of positive blood samples: after the positive blood sample is cracked and washed, the positive blood sample is centrifuged at 13000rpm for 5min, 100uL of supernatant is sucked and mixed with 100uL of sample treatment liquid uniformly, and then the positive blood sample is kept stand at room temperature for 10min. 200uL of liquid is sucked and slowly added into a sample adding hole of an immunochromatographic test strip, and the result is observed after 10 minutes;

(B) Colony specimen treatment: after a single separated colony is picked by a 1uL sampling rod and evenly mixed in 200uL sample treatment liquid, 200uL liquid is sucked and slowly added into an immunochromatographic test strip sample adding hole, and the result is observed after 10 min;

(C) Interpretation of the results: and developing by a quality control line, wherein the color development of the corresponding detection line represents a positive result, and the non-color development of the detection line represents a negative result. The quality control line does not develop color, and whether the detection line develops color or not, the test strip is invalid, and the result is invalid and needs to be detected again, as shown in fig. 4.

It will be understood by those skilled in the art that, unless otherwise defined, all terms (including technical and scientific terms) used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this utility model belongs. It will be further understood that terms, such as those defined in commonly used dictionaries, should be interpreted as having a meaning that is consistent with their meaning in the context of the prior art and will not be interpreted in an idealized or overly formal sense unless expressly so defined herein.

The meaning of "and/or" in the present utility model means that each exists alone or both exist.

"connected" as used herein means either a direct connection between components or an indirect connection between components via other components.

The protection of the present utility model is not limited to the above embodiments. Variations and advantages that would occur to one skilled in the art are included in the utility model without departing from the spirit and scope of the inventive concept, and the scope of the utility model is defined by the appended claims.

Claims (10)

1. The quick six-joint detection immunochromatography test strip for carbapenemase is characterized by mainly comprising a bottom plate, a partition plate, an NC film, a sample pad, a gold mark/milk mark pad and an absorption pad; the NC film is fixed on the bottom plate, the gold mark/milk mark pad and the absorption pad are respectively fixed at two ends of the NC film, the sample pad is lapped on the gold mark/milk mark pad, the partition plate is positioned in the middle of the NC film and is perpendicular to the NC film for fixation, and the left end of the partition plate is positioned at the joint of the sample pad and the gold mark/milk mark pad; three detection lines and a quality control line are respectively arranged on NC films at two sides of the partition board, monoclonal antibodies of the carbapenemases are respectively coated on the six detection lines, goat anti-mouse IgG antibodies are coated on the quality control line, and 3 kinds of monoclonal antibodies marked by colloidal gold/latex microspheres are respectively sprayed on gold/latex pads at two sides.

2. The rapid six-joint detection immunochromatographic strip for carbapenemase according to claim 1, wherein the monoclonal antibodies are a KPC protein monoclonal antibody, an NDM protein monoclonal antibody, a VIM protein monoclonal antibody, an IMP protein monoclonal antibody, an OXA-23 protein monoclonal antibody and an OXA-48 protein monoclonal antibody, respectively.

3. The rapid six-joint detection immunochromatographic test strip for carbapenemase according to claim 1, wherein the number of NC membranes is two, the two NC membranes are fixed on a bottom plate side by side, and the separator is positioned between the two NC membranes and is fixed perpendicular to the NC membranes.

4. The rapid six-joint detection immunochromatographic test strip for carbapenemase according to claim 3, wherein each NC membrane is independently provided with a gold label/milk label pad and an absorption pad.

5. The carbapenemase rapid six-joint detection immunochromatographic test strip of claim 1, wherein the test strip is sleeved with a plastic cartridge and is provided with a sample adding hole, a result observation window and an air hole.

6. The rapid six-joint detection immunochromatographic test strip for carbapenemase according to claim 5, wherein the number of result observation windows and ventilation holes is 2, and the result observation windows and ventilation holes are independently positioned on two sides of the partition board.

7. The carbapenemase rapid six-joint detection immunochromatographic test strip of claim 1, wherein the detection lines and the quality control lines are arranged at equal intervals, the widths are 5mm, and the intervals are 5mm.

8. The carbapenemase rapid six-joint detection immunochromatographic test strip of claim 1, wherein the colloidal gold/latex microsphere has a particle size of 20-40 nm.

9. The rapid six-joint detection immunochromatographic strip for carbapenemase according to claim 1, wherein the sample to be tested is an isolated single colony or a positive blood specimen.

10. The rapid six-joint detection immunochromatographic test strip for carbapenemase according to claim 1, wherein the bottom plate is a PVC bottom plate, the partition plate is a PVC partition plate, the NC film is Millipore135 in America, the sample pad is a glass cellulose film of Ahlstrom 8964 in Finland, the gold/milk mark pad is a polyester fiber film of Ahlstrom 6613 in Finland, and the absorption pad is cellulose filter paper.