CN109916893A - A kind of dry chemistry reagent piece of quantitative detection albumin content and preparation method thereof - Google Patents
A kind of dry chemistry reagent piece of quantitative detection albumin content and preparation method thereof Download PDFInfo
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Abstract
The present invention relates to clinical in vitro diagnosis in vitro reagent technique field, in particular to a kind of dry chemistry reagent piece of quantitative detection albumin content and preparation method thereof.The dry chemistry reagent piece includes four laminations, is followed successively by support layer, reagent layer, conversion zone and diffusion layer；Reagent in reagent layer includes buffer system, hydrophilic colloid, surfactant, anionic dye and cosolvent；Reagent in conversion zone includes hydrophilic colloid.Dry chemistry reagent piece testing result accuracy of the present invention is high, precision is good, specificity is good, and stability is good, long shelf-life, it is easy to operate, it is applied widely, be not necessarily to professional the advantages that, well supplement wet chemistry method shortcoming, its accuracy of measurement and precision are provided a great convenience already close to wet chemical determination method in clinical field of fast detection.
Description
Technical field
The present invention relates to clinical in vitro diagnosis in vitro reagent technique field, in particular to a kind of quantitative detection albumin content is done
Chemical reagent piece and preparation method thereof.
Background technique
Seralbumin is synthesized by liver parenchymal cell, its half-life period is about 15-19 days in blood plasma, is contained in blood plasma
A kind of protein that amount is most, molecule is minimum, solubility is big, function is more, accounts for the 40%-60% of Total plasma protein.It is by
The single chain protein matter of 585 amino acid composition, molecular weight is about 67kD, and mature albumin is one heart-shaped molecule, by 3
The similar α-helixstructure domain composition of structure.
The maintenance of plasma colloid osmotic pressure relies primarily on the albumin in blood plasma, and colloid osmotic pressure is between making vein end tissue
Liquid returns to the major impetus in blood return tube.Plasma albumin can be with the small organic molecule and inorganic ions of internal many slightly solubilities
The compound for forming ease of solubility is reversibly combined, types of transportation of these substances in blood circulation is become.Albumin is a kind of
With stiffness, collagenous substance, in the presence of heavy metal in human body ion, understand automatically in conjunction with heavy metal ion, thus by
Excretory system excretes, and plays the role of removing toxic substances.In addition, a kind of important nutriment in albumin or human body.White egg
White that metabolic turnover is also constantly carried out in blood plasma, plasma albumin decomposes the amino acid generated, can be used for synthetic tissue egg
White, oxygenolysis is with supplying energy or is transformed into other nitrogen substances.Furthermore albumin adjusts the metabolism of these hormones and drug
On, it is of great significance.
Albumin, which is generallyd use, at present is measured using liquid reagent in large-scale automatic clinical chemistry analyzer.Equipment itself compared with
For valuableness, do not have economy；It needs professional technician to operate instrument simultaneously, does not have due universality.Detect formality
Cumbersome, long flow path, waiting for a long time, analytic process is complex, brings the huge time to patient and bears.
Real-time test is an emerging technical field, mainly applies dry chemistry reagent, is easy to use and minimize.Mainly
Applied to emergency treatment, on-site test.Have many advantages, such as easy, quick, flexible and is operated without professional.It is domestic at present basic
For qualitative or semiquantitative determination, this dry chemistry reagent piece is by a kind of multilayer material such as filter paper, cellulose tablet, porous glue film
Whole reagents fixed in advance, the measurement range of linearity is little, not can avoid the interference of globulin transferrins and drug class.Quantitative inspection
The dry chemistry reagent card of survey is all external production.Using Johson & Johnson and company, Fuji as the multilayer dry slides method of representative, sense is utilized
The reagents such as reagent layer, auxiliary reagent layer, optical diffusion layer and distribution layer are coated in transparent support layer by the coating technology of ray film,
From the variation of the reverse side of support layer measurement reflectivity.Though above-mentioned dry chemistry reagent piece can quantitative detection, distribution layer or diffusion layer
Organic reagent need to be used, requires material installation high, production technology complexity, and very harmful to preparation personnel health, pollution
Environment.
Summary of the invention
In view of this, the present invention provides a kind of dry chemistry reagent piece of quantitative detection albumin content and its preparation sides
Method.The dry chemistry reagent piece testing result accuracy is high, and precision is good, and specificity is good, and stability is good, long shelf-life, operation letter
Just, applied widely.
In order to achieve the above-mentioned object of the invention, the present invention the following technical schemes are provided:
The present invention provides a kind of dry chemistry reagent pieces of quantitative detection albumin content, include four laminations, are followed successively by
Support layer, reagent layer, conversion zone and diffusion layer；
Reagent in reagent layer includes buffer system, hydrophilic colloid, surfactant, anionic dye and cosolvent；
Dosage of each component are as follows:
Reagent in conversion zone includes hydrophilic colloid.
Testing principle are as follows: liquid sample dropwise addition is evenly distributed in analoids diffusion layer, the high molecular weight protein class in sample
Penetration is into conversion zone, and the dyestuff in reagent layer can be continuously delivered into conversion zone, occurs with the albumin in sample aobvious
Colour response in the specific time, measures the variation of reflectivity under 630nm wavelength through reflective spectrophotometry, passes through standard song
The content of line computation albumin.Sample is added dropwise in diffusion layer, is measured in opposite one side, colored intensity is high, reduces sample-adding end and surveys
The raw bubble of fixed output quota, excess liq, turbidity and caused by interfere.Simultaneously because reagent layer and conversion zone are hydrophilic colloid, have
Very high colour developing evenness.Dry chemistry reagent piece of the present invention is used for the content of quantitative detection albumin, can be used for clinical quickly detection,
With good economic efficiency and application.
Dyestuff in reagent layer of the present invention can be continuously delivered into conversion zone, and detection, can reduce competitor in specific time
The interference of matter.The present invention uses hydrophilic filter membrane for diffusion layer, can make lamination is indivisible to become an entirety, and simplify system
Standby process.Dyestuff, which can be continuously delivered into conversion zone, carries out chromogenic reaction with albumin, and detection in specific time avoids endogenous
Property interference, realize fast and accurately quantitative analysis detect, improve detection accuracy, sensitivity.
Preferably, reagent layer is lower than conversion zone to the permeability of diffusible substance.
Preferably, buffer system is Tris buffer, succinate buffer, MOPS buffer or sweet ammonia in reagent layer
One or more of acid buffer, pH value is 4.0~4.5.
Preferably, buffer system is succinate buffer, and pH value is 4.0~4.15.
Preferably, surfactant be polyoxyethylene sorbitan monooleate 80, sorbitanmonolaureate,
Polyoxyethylene laurel ether, polyethylene glycol are to one or more of isooctyl phenyl ether.
Preferably, surfactant is polyoxyethylene laurel ether.
Preferably, anionic dye is bromocresol green or bromocresol purple.
Preferably, cosolvent is sodium chloride.
Preferably, dosage of each component in reagent layer are as follows:
Preferably, hydrophilic colloid is selected from polyvinyl alcohol, amylopectin, cellulose esters, agarose, polyvinylpyrrolidine
Ketone, polyacrylamide, hydroxypropyl methyl cellulose, polyethyleneimine, epoxy resin, polyurethane, methyl vinyl ether/maleic acid
One of anhydride copolymer is a variety of.
Preferably, hydrophilic colloid is cellulose esters in reagent layer.
Preferably, hydrophilic colloid is the mixture of cellulose esters and polyvinyl alcohol in conversion zone.
Preferably, the dosage of hydrophilic colloid is 50~150g/m in conversion zone2。
Preferably, diffusion layer is hydrophilic macromolecule filter membrane, hydrophilic macromolecule filter membrane is cellulose acetate, gathers inclined fluorine
The filter membrane of one or more of ethylene, polyether sulfone, polytetrafluoroethylene (PTFE) preparation, aperture are 0.45~8.0 micron.Diffusion layer can incite somebody to action
Sample is uniformly distributed while providing reflecting background.
Preferably, hydrophilic macromolecule filter membrane preferably 0.8~3.0 micron pore size.
Preferably, support layer is the polymer of polyethylene terephthalate, with a thickness of 50~300 microns.
Preferably, support layer is with a thickness of 100~200 microns.
The present invention also provides the preparation methods of the dry chemistry reagent piece, include the following steps:
Reagent solution reagent, reaction solution reagent are sequentially coated on support layer, it is dry, then after the drying by diffusion layer fitting
Conversion zone on, obtain successively include support layer, reagent layer, conversion zone and diffusion layer dry chemistry reagent piece；
Reagent in reagent layer includes buffer system, hydrophilic colloid, surfactant, anionic dye and cosolvent；
Dosage of each component are as follows:
Reagent in conversion zone includes hydrophilic colloid.
Preferably, the surface of support layer is by ultraviolet irradiation, corona or coating bottom layer treatment.
Preferably, the coating method of reagent layer and conversion zone includes spraying, levelling, hangs stream, rolls viscous or coating.
Preferably, the coating method of reagent layer is spraying, the coating method of conversion zone is coating.
Preferably, the mode of diffusion layer fitting is bonding, compacting.
The present invention provides dry chemistry reagent pieces of a kind of quantitative detection albumin content and preparation method thereof.The dry chemical
Analoids include four laminations, are followed successively by support layer, reagent layer, conversion zone and diffusion layer；Reagent in reagent layer includes buffering
System, hydrophilic colloid, surfactant, anionic dye and cosolvent；Dosage of each component are as follows: buffer system 0.01~
0.1mol/m2；5~50g/m of hydrophilic colloid2；0.5~5g/m of surfactant2；0.1~50mmol/m of anionic dye2；It helps
1~10mmol/m of solvent2；Reagent in conversion zone includes hydrophilic colloid.Compared with prior art, the present invention has following excellent
Point:
(1) operating process is simple, and pattern detection is directly added dropwise without preparing in reagent.
(2) the dry no moisture of analoids, analoids have good stability.
(3) result accuracy is high, can reduce endogenous interference.
(4) testing result is reproducible, can carry out quantitative detection, and the range of linearity is wide.
(5) preparation process is easy, and easy to operate, harmfulness, pollution are small.
Experiment shows that dry chemistry reagent piece testing result accuracy of the present invention is high, and precision is good, and specificity is good, surely
The advantages that qualitative good, long shelf-life is easy to operate, applied widely, is not necessarily to professional, well supplemented with wet chemistry method
Shortcoming, accuracy of measurement and precision are provided already close to wet chemical determination method in clinical field of fast detection
Great convenience.
Specific embodiment
The invention discloses dry chemistry reagent piece of a kind of quantitative detection albumin content and preparation method thereof, this field skills
Art personnel can use for reference present disclosure, be suitably modified realization of process parameters.In particular, it should be pointed out that all similar replacements and
Change apparent to those skilled in the art, they are considered as being included in the present invention.Method of the invention and
Using being described by preferred embodiment, related personnel can obviously not depart from the content of present invention, spirit and scope
It is interior that method described herein and application are modified or appropriate changes and combinations, carry out implementation and application the technology of the present invention.
In dry chemistry reagent piece of quantitative detection albumin content provided by the invention and preparation method thereof agents useful for same or
Instrument is available on the market.
Below with reference to embodiment, the present invention is further explained:
Embodiment 1: the preparation method of albumin dry chemistry reagent piece of the present invention
Transparent support layer material uses polyethylene terephthalate material, and with a thickness of 0.18mm, length 20cm is wide
Degree is 20cm.Surface is handled by ultraviolet irradiation, irradiation distance 10cm, and irradiation time 10 minutes.
The formula of reagent solution reagent is as follows:
The formula of reaction solution reagent is as follows:
Cellulose esters 15.4g/m2
Polyvinyl alcohol 95g/m2
Reagent solution is sprayed on the polyethylene terephthalate support layer by ultraviolet processing, is put into drying box and is dried
It is dry.Reaction solution is coated on the reagent layer of drying, is put into drying box and is dried.
It is 20cm, width 20cm by length, the cellulose acetate sheets purifying water-soaked that aperture is 0.8 micron shaves
Surface moisture is attached in the conversion zone of drying, and compacting is put into drying box and dries.
It is cut to the test-paper of 12mm × 12mm size with strip cutting machine, is sealed with aluminium foil bag.
20 μ L test samples are added dropwise in analoids the albumin sample for testing different content, with baffled photometer into
Row measurement, testing time are 3 minutes.The content of albumin in sample is calculated according to standard curve.
Embodiment 2: the accuracy analysis of the method for the invention
Test sample: 20 examinee's blood samples；
Contrasting detection method: with commercially available albumin detection reagent box (Bromocresol green) single liquid reagent in Hitachi 7080
It is detected on automatic clinical chemistry analyzer, 2.5 μ L samples is added by automatic clinical chemistry analyzer, 250 μ L reagents, 37 DEG C are reacted 2 points
Clock measures the absorbance value of 630nm, and the content of albumin in sample is calculated according to standard curve.
20 samples are measured respectively with 1 detection method of embodiment and contrasting detection method, and testing result is shown in Table 1.
Table 1 analyzes result
The results show that calculated R according to testing result2Value is 0.993, is greater than 0.95, shows the method for the invention
Testing result and contrast agent box testing result no significant difference have high accuracy (degree of conformity).
Embodiment 3: the Precision Analyze of the method for the invention
Test sample: any one blood serum sample；
Detection 10 times is repeated to same sample to be tested with 1 detection method of embodiment, testing result is shown in Table 2.
2 test sample albumin content of table (10 times) and standard rate (coefficient of variation)
The results show that standard rate is 1.74%, less than the 10% of standard requirements, show that the method for the invention has height
Precision.
Embodiment 4: the linear analysis of the method for the invention
Test sample: high-content albumin sample (60g/L)；
Albumin sample is diluted to 5 different contents, 0g/L, 15g/L, 30g/L, 45g/L, 60g/L is followed successively by, adopts
2 detections are carried out with each content of the detection method of embodiment 1 to above-mentioned sample, calculate coefficient R value, testing result is shown in
Table 3.
3 linear test result of table
The results show that calculating R value according to the testing result that 1 detection method of embodiment obtains is 0.9989, close to 1,
Show that the method for the invention has the good linearity in the range of 0.0g/L-60.0g/L.
Embodiment 5: the preparation method of albumin dry chemistry reagent piece of the present invention
Transparent support layer material uses polyethylene terephthalate material, with a thickness of 0.178mm, length 30cm,
Width is 20cm.Surface is handled by ultraviolet irradiation, irradiation distance 10cm, and irradiation time 60 minutes.
The formula of reagent solution reagent is as follows:
The formula of reaction solution reagent is as follows:
Polyvinyl alcohol 100g/m2
Reagent solution is sprayed on the polyethylene terephthalate support layer by ultraviolet processing, and reaction solution continuously coats
On reagent layer, it is put into drying box and dries.
It is 30cm, width 20cm by length, the teflon membrane filter surface spraying that aperture is 1.5 microns is uniform pure
Change water, be attached in the conversion zone of drying, is compacted, is put into drying box and dries.
It is cut to the test-paper of 12mm × 12mm size with strip cutting machine, is sealed with aluminium foil bag.
Embodiment 6: the stability analysis of the method for the invention
Condition of storage: sealing is placed in 2-8 DEG C of refrigerator.
Round of visits: when 2-8 DEG C 0, March, September, 15 months, 18 months.
It is detected with the detection method of embodiment 5 in above-mentioned round of visits, accuracy in computation, precision, linear model
It encloses, testing result is shown in Table 4.
4 stability test result of table
The results show that calculating R according to the testing result that 5 detection method of embodiment obtains2Value is all larger than 0.95, standard deviation
Rate is respectively less than 10%, the R value of standard requirements all close to 1, shows that the method for the invention stores 18 under the conditions of 2-8 DEG C
Month, analoids still have good accuracy, precision, the range of linearity.
Embodiment 7: the anti-Interference Analysis of the method for the invention
Test sample: control albumin standard (40g/L) compares and IgG globulin (26g/ is added in albumin standard
L)；
3 detections are carried out to above-mentioned sample respectively in 2,3,4,5,6min using the detection method of embodiment 5, are calculated real
Sample is tested relative to bias produced by check sample measurement result, testing result is shown in Table 5.
5 anti-interference test result of table
The results show that calculating the D in 3min according to the testing result that 5 detection method of embodiment obtainsobsLess than Dc, not
It observes that clinical significance influences, and reaches the content value of albumin standards, show the method for the invention in 3min
Fully reacting, and the IgG globulin for measuring 26g/L content does not significantly interfere with measurement result.
The preparation method of 1 other methods albumin dry chemistry reagent piece of comparative example
Transparent support layer material uses polyethylene terephthalate material, and with a thickness of 0.20mm, reagent layer is using poly-
Sulfone film, diffusion layer use nylon net cloth.Reagent layer is immersed in reagent solution, diffusion layer is immersed in diffusion liquid, is soaked completely
After take out, strike off.It is put into 30-50 DEG C of drying box, takes out, reagent layer is pasted on support layer, then be compacted expand after dry
Dissipate layer.
The formula of reagent solution reagent is as follows:
The formula of diffusion liquid reagent is as follows:
Triton X-100 10g/L
It is cut to the test-paper of 12mm × 12mm size with strip cutting machine, is sealed with aluminium foil bag.
20 μ L test samples are added dropwise in analoids the albumin sample for testing different content, with baffled photometer into
Row measurement, testing time are 3 minutes.The content of albumin in sample is calculated according to standard curve.
The Precision Analyze of 2 comparative example of comparative example, 1 method the method
Test sample: any one blood serum sample；
Detection 10 times is repeated to same sample to be tested with 1 detection method of comparative example, testing result is shown in Table 6.
6 test sample albumin content of table (10 times) and standard rate (coefficient of variation)
The results show that standard rate is 5.58%, hence it is evident that be greater than embodiment standard rate 1.74%, show of the present invention
Method has high precision.
The linear analysis of 3 comparative example of comparative example, 1 method the method
Test sample: high-content albumin sample (60g/L)；
Albumin sample is diluted to 5 different contents, 0g/L, 15g/L, 30g/L, 45g/L, 60g/L is followed successively by, adopts
2 detections are carried out with each content of the detection method of comparative example 1 to above-mentioned sample, calculate coefficient R value, testing result is shown in
Table 7.
7 linear test result of table
The results show that calculating R value according to the testing result that 1 detection method of comparative example obtains is 0.9901, and embodiment R
Value 0.9989 closer to 1 shows that the method for the invention has the good linearity in the range of 0.0g/L-60.0g/L.
The above is only a preferred embodiment of the present invention, it is noted that for the ordinary skill people of the art
For member, various improvements and modifications may be made without departing from the principle of the present invention, these improvements and modifications are also answered
It is considered as protection scope of the present invention.
Claims (10)
1. a kind of dry chemistry reagent piece of quantitative detection albumin content, which is characterized in that include four laminations, be followed successively by support
Layer, reagent layer, conversion zone and diffusion layer；
Reagent in the reagent layer includes buffer system, hydrophilic colloid, surfactant, anionic dye and cosolvent；
Dosage of each component are as follows:
Reagent in the conversion zone includes hydrophilic colloid.
2. dry chemistry reagent piece according to claim 1, which is characterized in that in the reagent layer, buffer system Tris
One or more of buffer, succinate buffer, MOPS buffer or glycine buffer, pH value is 4.0~4.5；Table
Face activating agent is polyoxyethylene sorbitan monooleate 80, sorbitanmonolaureate, polyoxyethylene laurel ether, poly- second
Glycol is to one or more of isooctyl phenyl ether；Anionic dye is bromocresol green or bromocresol purple；Cosolvent is chlorination
Sodium.
3. dry chemistry reagent piece according to claim 1, which is characterized in that dosage of each component in the reagent layer are as follows:
4. dry chemistry reagent piece according to claim 1, which is characterized in that the hydrophilic colloid be selected from polyvinyl alcohol,
Amylopectin, cellulose esters, agarose, polyvinylpyrrolidone, polyacrylamide, hydroxypropyl methyl cellulose, polyethyleneimine
One of amine, epoxy resin, polyurethane, Copolymer of Methyl Vinyl Ether/Maleic Anhydride are a variety of.
5. dry chemistry reagent piece according to claim 1, which is characterized in that the dosage of hydrophilic colloid in the conversion zone
For 50~150g/m2。
6. dry chemistry reagent piece according to claim 1, which is characterized in that the diffusion layer is hydrophilic macromolecule filter
Film, the hydrophilic macromolecule filter membrane are one of cellulose acetate, Kynoar, polyether sulfone, polytetrafluoroethylene (PTFE) or several
Filter membrane prepared by kind, aperture are 0.45~8.0 micron.
7. dry chemistry reagent piece according to any one of claim 1 to 6, which is characterized in that the support layer is poly- pair
The polymer of ethylene terephthalate, with a thickness of 50~300 microns.
8. the preparation method of dry chemistry reagent piece as described in any one of claims 1 to 7, which is characterized in that including walking as follows
It is rapid:
Reagent solution reagent, reaction solution reagent are sequentially coated on support layer, it is dry, then by diffusion layer fitting after the drying anti-
Answer on layer, obtain successively include support layer, reagent layer, conversion zone and diffusion layer dry chemistry reagent piece；
Reagent in the reagent layer includes buffer system, hydrophilic colloid, surfactant, anionic dye and cosolvent；
Dosage of each component are as follows:
Reagent in the conversion zone includes hydrophilic colloid.
9. preparation method according to claim 8, which is characterized in that ultraviolet irradiation, electricity are passed through in the surface of the support layer
Dizzy or coating bottom layer treatment.
10. preparation method according to claim 8 or claim 9, which is characterized in that the coating method of the reagent layer and conversion zone
Comprising spraying, levelling, hangs and flow, roll viscous or be coated with.
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| CN113092781A (en) * | 2021-03-31 | 2021-07-09 | 深圳市锦瑞生物科技有限公司 | Preparation method of albumin determination reagent ball, reagent ball and microfluidic chip |
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| CN111879591A (en) * | 2020-09-01 | 2020-11-03 | 中科嘉睿(天津)医疗科技发展有限公司 | Dry chemical standard reagent tablet and preparation method thereof |
| CN113092781A (en) * | 2021-03-31 | 2021-07-09 | 深圳市锦瑞生物科技有限公司 | Preparation method of albumin determination reagent ball, reagent ball and microfluidic chip |
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Application publication date: 20190621 |