CN113009150A - Concentration device for collecting urine microalbumin in sweat, detection kit comprising same and application thereof - Google Patents
Concentration device for collecting urine microalbumin in sweat, detection kit comprising same and application thereof Download PDFInfo
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- CN113009150A CN113009150A CN202110194222.1A CN202110194222A CN113009150A CN 113009150 A CN113009150 A CN 113009150A CN 202110194222 A CN202110194222 A CN 202110194222A CN 113009150 A CN113009150 A CN 113009150A
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
- G01N33/6803—General methods of protein analysis not limited to specific proteins or families of proteins
- G01N33/6827—Total protein determination, e.g. albumin in urine
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/38—Diluting, dispersing or mixing samples
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/40—Concentrating samples
- G01N1/4005—Concentrating samples by transferring a selected component through a membrane
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/40—Concentrating samples
- G01N1/4044—Concentrating samples by chemical techniques; Digestion; Chemical decomposition
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/543—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals
- G01N33/54313—Immunoassay; Biospecific binding assay; Materials therefor with an insoluble carrier for immobilising immunochemicals the carrier being characterised by its particulate form
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/558—Immunoassay; Biospecific binding assay; Materials therefor using diffusion or migration of antigen or antibody
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/577—Immunoassay; Biospecific binding assay; Materials therefor involving monoclonal antibodies binding reaction mechanisms characterised by the use of monoclonal antibodies; monoclonal antibodies per se are classified with their corresponding antigens
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- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/40—Concentrating samples
- G01N1/4005—Concentrating samples by transferring a selected component through a membrane
- G01N2001/4016—Concentrating samples by transferring a selected component through a membrane being a selective membrane, e.g. dialysis or osmosis
Abstract
The invention relates to a concentration device for collecting urine microalbumin in sweat, a detection kit comprising the same and application of the concentration device. According to the kit for detecting the urine microalbumin in sweat, after the sample is added with the urine microalbumin immunochromatography detection card after reaction and concentration, the sample can be detected without waiting for a long time, the operation is simple, the sample processing time is short, the time cost is reduced, and the kit is suitable for clinical rapid diagnosis and detection.
Description
Technical Field
The invention belongs to the field of in-vitro diagnosis, and relates to a concentration device for collecting urine microalbumin in sweat, a detection kit comprising the same and application thereof.
Background
The albumin accounts for 60% of the total amount of plasma, has a molecular weight of 69kD, and is a macromolecular protein with negative charges. Normally only a very small amount of albumin is excreted by urine to the outside of the body. The glomerular capillary basement membrane has a filtering function, and the diameter of a membrane hole is 5.5 nm. The radius of albumin is 3.6nm, and albumin is difficult to pass through glomerular basement membrane under normal state. Any pathology that causes increased permeability of the glomerular basement membrane can lead to the excretion of albumin. Normally, urine contains very little albumin, less than 20mg/L, and is called Microalbumin (MAU). If the microalbuminuria in urine is in the range of 20mg/L-200mg/L, the microalbuminuria belongs to the critical stage of urine microalbuminuria, namely, the critical stage of urine microalbuminuria which exceeds the upper limit of normal reference of healthy people and is not detected. Urine microalbumin can be caused in early stages of primary kidney diseases, hypertension and diabetes combined with kidney injury. The detection of urine trace protein is the most sensitive and reliable diagnostic index for early detection of renal injury. Regular detection of microalbumin in urine should be done once a year by the average person, and every 3 months by the increased patients, which has a positive effect on prevention and early treatment of renal disease. The monitoring of urine microalbumin can be used as an index for early diagnosis and long-term follow-up of various primary and secondary kidney injuries.
At present, most of the research on sweat collecting devices is directed, but because the amount of sweat is small, if direct detection is carried out, the sensitivity may be reduced due to low concentration, and the human health is not easy to monitor in time. It is therefore desirable to develop a device that can concentrate and accelerate the reaction after sweat collection to improve sensitivity.
Disclosure of Invention
In view of the above, the main objective of the present invention is to provide a concentration device for collecting urine microalbumin in sweat, a detection kit comprising the same, and applications thereof.
In order to achieve the purpose, the invention provides a concentration device for collecting urine microalbumin in sweat, which comprises a tube body, wherein a hydrophobic membrane and a marking membrane are respectively arranged in the tube body from bottom to top, a rubber pad is arranged at the bottom outside the tube body, a vacuum pump is connected below the outside of the tube body, the vacuum pump is connected with a needle head, and the needle head is connected with the bottom of the tube body.
In a specific embodiment of the present invention, the hydrophobic membrane and the marking membrane are respectively clamped on the inner wall of the tube body; set up like this because during the centrifugation, can throw the waste liquid into the waste liquid storehouse through hydrophobic membrane after buffer solution and the abundant reaction of sample, and the liquid after the concentration is retained at the socle portion.
In a specific embodiment of the present invention, the tube contains a buffer solution, the labeling membrane is soaked in the buffer solution, and the size of the labeling membrane is smaller than or equal to the inner diameter of the tube; so that the labeling membrane is completely soaked in the buffer solution, and the antibody on the labeling membrane can fully react with sweat.
In one embodiment of the present invention, wherein the hydrophobic membrane has a pore size of less than 200nm, the arrangement is to prevent the buffer from infiltrating in the reaction stage, and the pore size of less than 200nm is to prevent the labeled antibody from infiltrating into the stepped reaction chamber because the diameter of the labeled fluorescent microsphere is greater than 200. (ii) a Specifically, the hydrophobic membrane is an ML-QF series amphiphobic microporous membrane, the material is polyethylene terephthalate (PET), the pore diameter is 10 micrometers, and the membrane thickness is 70-100 micrometers.
In one embodiment of the invention, the volume ratio of the pipe body to the vacuum pump is (1-10) to (0.9-1.9); therefore, the optimal concentration effect can be achieved without wasting raw materials after optimization.
In a specific embodiment of the present invention, the needle is respectively inserted through the rubber pad and the bottom of the tube, and the other end of the needle is lower than the height of the hydrophobic membrane; the design is designed to ensure that the hydrophobic membrane cannot be broken and the liquid after reaction can be sucked.
In a specific embodiment of the present invention, the tube body has a hollow cylindrical structure or a rectangular parallelepiped structure.
In a specific scheme of the invention, a mouse anti-human urinary microalbumin monoclonal antibody marked by latex fluorescent microspheres and a rabbit IgG polyclonal antibody marked by the latex fluorescent microspheres are marked on the marking membrane; the molar ratio of the mouse anti-human urine microalbumin monoclonal antibody marked by the latex fluorescent microspheres to the rabbit IgG polyclonal antibody marked by the latex fluorescent microspheres is 1: 0.2-4, preferably 1:1.2, so that the sensitivity and specificity of the detection kit for urine Microalbumin (MAU) in liquid can be improved.
In order to achieve the above object, the present invention further provides a kit for detecting urine microalbumin in sweat, the kit comprising the above concentration device and a urine microalbumin immunochromatography detection card.
In a specific embodiment of the invention, the urine microalbumin immunochromatography detection card comprises a test strip, the test strip comprises a detection line and a quality control line, the detection line is coated with another urine Microalbumin (MAU) monoclonal antibody, and the quality control line is coated with a goat anti-rabbit polyclonal antibody; the test strip further comprises a PVC plate, wherein a sample adding pad, a coating pad and a water absorption pad which are connected in sequence are fixed on the PVC plate, and a detection line and a quality control line are arranged on the coating pad in sequence.
In a specific embodiment of the present invention, the membrane-scratching concentration ratio of the another monoclonal antibody against mouse human urine microalbumin to the polyclonal antibody against goat rabbit is (1-2): (1-2), preferably 1: 1; thus, the raw materials are not wasted after the optimization, and the measurement result is relatively accurate.
In a specific embodiment of the present invention, the urine microalbumin immunochromatography test card further comprises a card shell for holding a test strip; the card case includes:
a bottom trough connected to the PVC sheet;
the upper cover is connected with the bottom groove, and a sample adding hole for adding samples to the sample adding pad is arranged on the upper cover;
and the observation window is arranged on the upper cover and is used for data acquisition of the detection line and the quality control line.
In order to achieve the above object, the present invention also provides a method for using a kit for detecting urinary Microalbumin (MAU) in sweat, comprising the steps of:
1) mixing the sweat and the buffer solution according to the proportion of 1-10: 10;
2) and (2) dripping 30-100 mu L of the sample diluted in the step 1) into a sample adding hole of the sweat microalbumin-urine detection kit, standing at room temperature for 5-20 min, inserting the sweat microalbumin-urine detection kit into a fluorescence immunoassay analyzer for detection, and immediately obtaining a detection result.
In a specific embodiment of the present invention, the sweat collection is prior art, and need not be described herein; for example, sweat may be collected by a flexible sweat collector.
In one embodiment of the present invention, the sweat sample is selected from a part which is easy to generate sweat and convenient to collect, such as forehead, neck or underarm.
The application has the specific operation process and principle that: adding sweat collected by a flexible sweat collector into a concentration device for collecting urine microalbumin in the sweat by the prior art, adding a buffer solution, uniformly mixing (the sweat reacts with an antibody marked on a marking membrane), downwards pumping liquid by a vacuum pump, controlling the volume absorbed by the vacuum pump at the lowest part of the concentration device for collecting the urine microalbumin in the sweat and the molecular diameter of a hydrophobic membrane, pumping small-molecule water to the hydrophobic membrane under negative pressure, keeping the large-molecule fluorescent microsphere-marked antibody on the upper layer of the hydrophobic membrane, and discharging redundant water to a waste liquid collection device to perform a concentration function.
Compared with the prior art, the invention has the following beneficial effects:
1. the concentration device for collecting urine Microalbumin (MAU) in sweat can be matched with a flexible wearable sweat collection device to be used for detecting the MAU in the concentrated sweat and quantitatively measuring the content of the urine microalbumin in the human sweat in vitro; the sensitivity is increased after concentration, and the detection efficiency is greatly improved.
2. The kit for detecting the urine Microalbumin (MAU) in the sweat has the advantages of high sensitivity, strong stability, excellent accuracy and precision.
3. According to the kit for detecting the urine microalbumin in sweat (MAU), after a sample is reacted and concentrated and added with the urine microalbumin immunochromatography detection card, the sample can be detected without waiting for a long time, the operation is simple, the sample processing time is short, the time cost is reduced, and the kit is suitable for clinical rapid diagnosis and detection.
4. The kit for detecting the urine Microalbumin (MAU) in the sweat disclosed by the invention is characterized in that an antibody marked by fluorescent microspheres is added into a concentration device in advance, so that the reaction is fully carried out.
Drawings
FIG. 1 is a schematic diagram of a concentration device for collecting urine Microalbumin (MAU) in sweat according to the present invention;
FIG. 2 is a schematic structural diagram of the MAU immunochromatographic detection card provided in the present invention;
FIG. 3A is a schematic diagram of the internal structure of an upper cover of the MAU immunochromatographic assay card provided in the present invention;
FIG. 3B is a schematic diagram of the internal structure of a bottom slot of the MAU immunochromatographic assay card provided in the present invention;
FIG. 4 is a schematic diagram showing a standard curve of MAU in example 1 of the present invention;
wherein, 1 a-tube body, 2 a-marking film, 3 a-hydrophobic film, 4 a-needle head, 5 a-vacuum pump; 6 a-rubber pad, 1-PVC board, 2-coating pad, 3-marker pad, 4-absorbent pad, 5-detection line, 6-quality control line, 7-marker junction, 8-sample, 9-sample adding pad, 11-upper cover, 12-bottom groove, 13-sample adding hole, 14-observation window, 15-test strip placing area, 16-positioning column, 17-positioning hole, 18-first limiting part, 19-second limiting part and 20-third limiting part.
Detailed Description
The invention is further described below with reference to the accompanying drawings.
The following materials or reagents, unless otherwise specified, are commercially available.
Example 1 preparation of urine Microalbumin (MAU) immunochromatography assay kit
1. Preparation of urine Microalbumin (MAU) immunochromatography detection kit
1) Concentration device for collecting urine Microalbumin (MAU) in sweat
As shown in figure 1, a hydrophobic membrane 3a (with the aperture of 150 nm; concretely, the hydrophobic membrane is an ML-QF series double-hydrophobic microporous membrane made of polyethylene terephthalate (PET), the aperture of 10 microns and the membrane thickness of 70-100 microns) and a marking membrane 2a are respectively clamped on the inner wall of a tube body 1a from bottom to top, meanwhile, a rubber pad 6a is arranged at the bottom outside the tube body 1a, and then, the bottom outside the tube body 1a is connected with a vacuum pump 5a through a needle 4a (with the inner diameter of 1 mm); the size of the marking film 2a is smaller than or equal to the inner diameter of the tube body 1a, for example, the size of the marking film 2a may be 90mm, and the inner diameter of the tube body 1a may be 1 cm. The volume ratio of the tube body 1a to the vacuum pump 5a is (1-10): 0.9-1.9), for example, the volume of the tube body 1a is 1.5ml (diameter 1cm), and the volume of the vacuum pump is 125cm3(5cm by 5 cm). The needle 4a penetrates through the rubber pad 6s and the bottom of the tube body 1a to enter the tube body 1a, and the other end of the needle 1a is lower than the hydrophobic membrane 3 a. The pipe body 1a is of a hollow cylindrical structure. A mouse anti-human MYO monoclonal antibody marked by latex fluorescent microspheres and a rabbit IgG polyclonal antibody marked by the latex fluorescent microspheres are marked on the marking film 2 a; the molar ratio of the mouse anti-human MYO monoclonal antibody marked by the latex fluorescent microsphere to the rabbit IgG polyclonal antibody marked by the latex fluorescent microsphere is 1: 0.2-4, and preferably 1: 1; the marking membrane 2a is a nitrocellulose membrane.
2) Fluorescent microsphere labeled mouse anti-human urine Microalbumin (MAU) monoclonal antibody
Taking 0.1g of latex fluorescent microspheres, centrifuging for 30min by using a centrifuge (the rotating speed is 10000r/min, 20mL at most once), discarding supernatant after centrifugation, redissolving by using 10mL of marking buffer solution, adding 2mg of urine Microalbumin (MAU) marking antibody, uniformly mixing, adding 10mg of carbodiimide, stirring and reacting for 1h at room temperature, centrifuging for 15min (the rotating speed is 10000r/min), discarding supernatant, diluting by using 20mL of marker diluent, redissolving, and uniformly mixing for later use.
3) Fluorescent microsphere labeled rabbit IgG polyclonal antibody
Taking 0.05g of latex fluorescent microspheres, centrifuging for 30min by using a centrifuge (the rotating speed is 10000r/min for one time and is at most 20mL), discarding the supernatant after centrifugation, redissolving by using 5mL of marking buffer solution, adding 1mg of rabbit IgG, mixing uniformly, adding 5mg of carbodiimide, stirring and reacting for 1h at room temperature, centrifuging for 15min (the rotating speed is 10000r/min), discarding the supernatant, diluting by using 10mL of marking substance diluent, redissolving, and mixing uniformly for later use.
4) Preparation of detection line and quality control line
The other MAU-coated antibody and goat anti-rabbit polyclonal antibody were diluted with phosphate buffer (pH 7.4) and streaked on nitrocellulose membrane 2(NC membrane) at a concentration of 1mg/mL using a streaking au-spraying instrument. The detection line 5(T line) containing another MAU coated antibody and the quality control line 6(C line) containing goat anti-rabbit polyclonal antibody are dried for 4 hours at 37 ℃ and the humidity of less than 30 percent to prepare the marker pad 3.
5) Assembly of immunochromatography test card
Firstly, a nitrocellulose membrane 2 is bonded on a PVC (polyvinyl chloride) plate 1, then, one end close to a quality control line 6 on the nitrocellulose membrane 2 is lapped with a water absorption pad 4, one end close to a detection line 5 of the nitrocellulose membrane 2 is lapped with a sample adding pad 9, a strip cutting machine is used for cutting the test strip into test strips with the thickness of 4mm +/-0.1 mm, and the test strips are put into a card shell to prepare the MAU (seen in figure 2) immunochromatography detection card.
The card housing is selected from the prior art, for example, the card housing (as shown in fig. 3A and 3B) may include: a bottom tank 12 connected to the PVC plate 1; an upper cover 11 connected to the bottom tank 12, the upper cover 11 being provided with a sample application hole 13 for applying a sample to the sample application pad 9; and the observation window 14 is arranged on the upper cover 11 and is used for data acquisition of the detection line 5 and the quality control line 6.
As shown in fig. 3B, the bottom tank 12 includes: a plurality of positioning holes 17 which are symmetrically distributed and positioned on the inner surface of the test strip, wherein a plurality of first limiting parts 18 used for limiting the test strip to move transversely and second limiting parts 19 used for limiting the test strip to move longitudinally are arranged among the plurality of positioning holes 17; the first limiting part 18 and the second limiting part 19 which are symmetrically arranged enclose a paper strip placing area 15 (a dotted line area) for placing the test paper strip;
as shown in fig. 3A, the upper cover 11 includes: a plurality of positioning posts 16 which are matched with a plurality of positioning holes 17, and are matched with each other to fix the upper cover 11 and the bottom groove 12 together; the upper cover 11 further includes a third limiting portion 20 for limiting the up and down movement of the test strip.
An observation window 14 for data acquisition is arranged above the coated pad 2 to expose all the detection lines 5 and the quality control lines 6 for collecting detection results; and the observation window 14 is arranged on the upper cover 11 at a position corresponding to the middle part of the test strip placement area 15. The upper cover 11 is provided with a sample adding hole at a position corresponding to the sample adding pad 9 for dropping the sample 8 on the sample adding pad 9. The distance between the detection line and the sample adding hole is 15-25 mm.
2. Preparation of buffer
5.79g of sodium citrate, 9.54g of trisodium citrate, 1g of Proclin300 and 2g of bovine serum albumin were added to a beaker, stirred while adding water until the sodium citrate, the trisodium citrate, Proclin300 and bovine serum albumin were dissolved completely, and then the volume of the solution was determined to 1000mL with water, thereby obtaining a buffer solution containing 0.06M of sodium citrate, 0.06M of trisodium citrate, 0.1% (v/v) of Proclin300 and 0.2 wt% of bovine serum albumin for use.
3. Detection of
1) Adding the collected sweat into a tube body 1a of the concentration device for collecting the urine microalbumin in the sweat, and mixing the sweat with a buffer solution according to the volume ratio of 1:1 to obtain a sample mixed solution (a concentrated sample); then, the sample is extracted by a vacuum pump 5a of the concentration device of embodiment 1 of the present invention to obtain a sample mixture (concentrated sample);
2) and (2) taking 60 mu L of the sample mixed solution obtained in the step 1), dropwise adding the sample mixed solution into a sample adding hole of the urine microalbumin immunochromatography detection card, standing at room temperature for 15min, and then inserting the urine microalbumin immunochromatography detection card into a fluorescence immunoassay analyzer for detection to obtain a detection result immediately.
The detection principle is as follows: when the concentrated sweat buffer solution to be detected is added to the sample adding pad 9, the antigen will mix with the marker for reaction and will carry out chromatography along the nitrocellulose membrane 2, and will react with the detection line 5 and the quality control line 6 respectively. When the test result is valid, the quality control line 6 displays a certain light intensity. At this time, the ratio (T/C) of the light signal intensity on the detection line 5 to the light signal intensity of the quality control line 6 is in positive correlation with the sample concentration, and the concentration of the sample to be detected can be obtained through calculation of a standard curve.
4. Establishment of a Standard Curve
MAU standard substances (0, 10, 25, 50, 150 and 500ng/mL) with different concentrations are prepared, the MAU kit prepared in the invention is used for detection, the detection sensitivity is 10ng/mL, the detection range is 10-500 ng/mL, curve fitting is carried out by adopting Logistic four parameters, and a standard curve is shown in FIG. 4. Table 1 shows the results of the measured T/C values of the standards at different concentrations, and FIG. 4 shows a four-parameter curve fitting, R2As can be seen from the data in table 1 and fig. 2, the MAU has a better standard curve fit at 0.99995.
TABLE 1 test data
| Standard concentration ng/mL | 0 | 10 | 25 | 50 | 150 | 500 |
| T/C value | 0 | 0.051 | 0.119 | 0.25 | 0.719 | 1.498 |
Example 2 comparison of measured values with clinical diagnostic information
The results obtained using the buffer and concentration device and MAU immunochromatographic test card prepared in example 1 were compared with clinical diagnostic information. By collecting 40 sweat samples, with 22 total normal persons; the number of patients was 18. 18 sweat samples clinically diagnosed as impaired renal function were obtained, and the results are shown in table 2 below.
TABLE 2
As can be seen from the test results in table 2, the comparison result between the test result of the concentration device for collecting serum amyloid a in sweat and the MAU immunochromatography test card of the present invention and the clinical diagnosis information is as follows:
sensitivity is 94.70% (95% CI 75.36% -99.06%),
specificity 88.89% (95% CI 79.76% -96.90%)
Therefore, the concentration device of the invention can be combined with the MAU immunochromatography detection card to detect sweat to assist the clinical diagnosis.
20 samples (samples No. 1-20 in Table 2) were selected and concentrated (concentration ratio 10 times) and directly measured by the concentration device of the embodiment 1 of the present invention, and the results were compared with the clinical diagnosis results, as shown in Table 3; the results in Table 3 show that the values measured without concentration are overall lower than those measured after concentration. This indicates that after concentration, the measured results are substantially greater than before concentration, indicating that concentration further affects the measured results.
TABLE 3
The above-mentioned embodiments are merely examples provided to fully illustrate the present invention, and the scope of the present invention is not limited thereto. The equivalent substitution or change made by the technical personnel in the technical field on the basis of the invention is all within the protection scope of the invention. The protection scope of the invention is subject to the claims.
Claims (10)
1. The utility model provides a enrichment facility for collecting urine microalbumin in sweat, its characterized in that, enrichment facility includes the body, be equipped with hydrophobic membrane and mark membrane respectively from bottom to top in the body, the outside bottom of body is equipped with the rubber pad, the outside below of body is connected with the vacuum pump, vacuum pump connection has the syringe needle, the syringe needle with the bottom of body is connected.
2. The concentrating device for collecting urinary microalbumin in sweat of claim 1, wherein the hydrophobic membrane and the marking membrane are respectively clamped on the inner wall of the tube; buffer solution is contained in the tube body, the marking film is soaked in the buffer solution, and the size of the marking film is smaller than or equal to the inner diameter of the tube body.
3. The concentrating device for collecting urinary microalbumin in sweat of claim 1, wherein the hydrophobic membrane has a pore size of less than 200 nm; the volume ratio of the pipe body to the vacuum pump is (1-10) to (0.9-1.9).
4. The concentrating device for collecting urinary microalbumin in sweat of claim 1, wherein the needle is inserted through the rubber pad and the bottom of the tube, and the other end of the needle is lower than the height of the hydrophobic membrane; the tube body is of a hollow cylindrical structure or a rectangular structure.
5. The concentrating device for collecting urinary microalbumin in sweat of claim 1, wherein the labeled membrane is labeled with a mouse anti-human urinary microalbumin monoclonal antibody labeled with latex fluorescent microspheres, a rabbit IgG polyclonal antibody labeled with latex fluorescent microspheres; the molar ratio of the mouse anti-human urinary microalbumin monoclonal antibody marked by the latex fluorescent microspheres to the rabbit IgG polyclonal antibody marked by the latex fluorescent microspheres is 1: 0.2-4; the marking membrane is a nitrocellulose membrane.
6. A kit for detecting urinary microalbumin in sweat, comprising the concentration device of any one of claims 1 to 5 and a urinary microalbumin immunochromatography test card.
7. The kit of claim 6, wherein the microalbumin immunochromatography test card comprises a strip of paper containing a test line coated with another monoclonal antibody against microalbumin and a quality control line coated with a polyclonal antibody against goat rabbit; the test strip further comprises a PVC plate, wherein a sample adding pad, a coating pad and a water absorption pad which are connected in sequence are fixed on the PVC plate, and a detection line and a quality control line are arranged on the coating pad in sequence.
8. The kit of claim 6, wherein the another strain of mouse anti-human urinary microalbumin monoclonal antibody to goat anti-rabbit polyclonal antibody has a cross-membrane concentration ratio of (1-2): (1-2).
9. The kit for detecting urinary microalbumin in sweat of claim 6, wherein the urine microalbumin immunochromatography test card further comprises a card case for holding a test strip; the card case includes:
a bottom trough connected to the PVC sheet;
the upper cover is connected with the bottom groove, and a sample adding hole for adding samples to the sample adding pad is arranged on the upper cover;
and the observation window is arranged on the upper cover and is used for data acquisition of the detection line and the quality control line.
10. A method of using the urine microalbumin in sweat detection kit of any one of claims 6 to 9, comprising the steps of:
1) mixing the sweat and the buffer solution according to the proportion of 1-10: 10;
2) dripping 30-100 mu L of the sample diluted in the step 1) into a sample adding hole of a detection kit for the urine microalbumin in sweat, standing at room temperature for 5-20 min, inserting the detection kit for the urine microalbumin in sweat into a fluorescence immunoassay analyzer for detection, and immediately obtaining a detection result;
the sweat sample is selected from forehead, neck or underarm.
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| Application Number | Priority Date | Filing Date | Title |
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| CN202110194222.1A CN113009150A (en) | 2021-02-20 | 2021-02-20 | Concentration device for collecting urine microalbumin in sweat, detection kit comprising same and application thereof |
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| CN202110194222.1A CN113009150A (en) | 2021-02-20 | 2021-02-20 | Concentration device for collecting urine microalbumin in sweat, detection kit comprising same and application thereof |
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| CN202110194222.1A Pending CN113009150A (en) | 2021-02-20 | 2021-02-20 | Concentration device for collecting urine microalbumin in sweat, detection kit comprising same and application thereof |
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Citations (11)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102749443A (en) * | 2011-04-22 | 2012-10-24 | 国家纳米科学中心 | Double layer micro fluidic chip device and its application in immunodetection |
| CN103063834A (en) * | 2012-12-28 | 2013-04-24 | 三诺生物传感股份有限公司 | Method and system for analysis of immune quantitative chromatographic assay strip |
| CN103323590A (en) * | 2013-06-08 | 2013-09-25 | 上海云泽生物科技有限公司 | Quantitative detection device based on fibrous-membrane gathering and separation and detection method thereof |
| CN106188300A (en) * | 2015-05-07 | 2016-12-07 | 温州市人民医院 | A kind of method for quickly purifying keeping bladder chalone C antibody-latex covalent coupling complex activity |
| CN106596968A (en) * | 2016-08-23 | 2017-04-26 | 广东优尼德生物科技有限公司 | Dot gold infiltration kit for detecting urine microalbumin, and application of dot gold infiltration kit |
| CN108106922A (en) * | 2018-01-31 | 2018-06-01 | 迈克医疗电子有限公司 | A kind of concentration mixing mechanism |
| CN109847816A (en) * | 2019-01-03 | 2019-06-07 | 温州医科大学 | A kind of microfluidic separation chip |
| CN110488006A (en) * | 2019-09-26 | 2019-11-22 | 天津华科泰生物技术有限公司 | A kind of immunochromatographydetection detection card and preparation method thereof of 3 sample albumen 1 of quick detection chitinase |
| CN111077325A (en) * | 2018-10-19 | 2020-04-28 | 上海快灵生物科技有限公司 | Microporous membrane interception and aggregation biochemical detection device and detection method thereof |
| CN111398593A (en) * | 2020-04-03 | 2020-07-10 | 天津华科泰生物技术有限公司 | Rapid combined detection card and preparation method and application thereof |
| CN111398589A (en) * | 2020-02-13 | 2020-07-10 | 北京华科泰生物技术股份有限公司 | Immunochromatography kit for rapidly detecting novel coronavirus N protein and preparation method and application thereof |
-
2021
- 2021-02-20 CN CN202110194222.1A patent/CN113009150A/en active Pending
Patent Citations (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102749443A (en) * | 2011-04-22 | 2012-10-24 | 国家纳米科学中心 | Double layer micro fluidic chip device and its application in immunodetection |
| CN103063834A (en) * | 2012-12-28 | 2013-04-24 | 三诺生物传感股份有限公司 | Method and system for analysis of immune quantitative chromatographic assay strip |
| CN103323590A (en) * | 2013-06-08 | 2013-09-25 | 上海云泽生物科技有限公司 | Quantitative detection device based on fibrous-membrane gathering and separation and detection method thereof |
| CN106188300A (en) * | 2015-05-07 | 2016-12-07 | 温州市人民医院 | A kind of method for quickly purifying keeping bladder chalone C antibody-latex covalent coupling complex activity |
| CN106596968A (en) * | 2016-08-23 | 2017-04-26 | 广东优尼德生物科技有限公司 | Dot gold infiltration kit for detecting urine microalbumin, and application of dot gold infiltration kit |
| CN108106922A (en) * | 2018-01-31 | 2018-06-01 | 迈克医疗电子有限公司 | A kind of concentration mixing mechanism |
| CN111077325A (en) * | 2018-10-19 | 2020-04-28 | 上海快灵生物科技有限公司 | Microporous membrane interception and aggregation biochemical detection device and detection method thereof |
| CN109847816A (en) * | 2019-01-03 | 2019-06-07 | 温州医科大学 | A kind of microfluidic separation chip |
| CN110488006A (en) * | 2019-09-26 | 2019-11-22 | 天津华科泰生物技术有限公司 | A kind of immunochromatographydetection detection card and preparation method thereof of 3 sample albumen 1 of quick detection chitinase |
| CN111398589A (en) * | 2020-02-13 | 2020-07-10 | 北京华科泰生物技术股份有限公司 | Immunochromatography kit for rapidly detecting novel coronavirus N protein and preparation method and application thereof |
| CN111398588A (en) * | 2020-02-13 | 2020-07-10 | 北京华科泰生物技术股份有限公司 | Use method of immunochromatography kit for rapidly detecting novel coronavirus N protein |
| CN111398593A (en) * | 2020-04-03 | 2020-07-10 | 天津华科泰生物技术有限公司 | Rapid combined detection card and preparation method and application thereof |
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