CN103055967B - A kind of preparation of simple to operate, low cost, multichannel micro-fluidic chemiluminescence paper chip and the application in detecting at the scene - Google Patents
A kind of preparation of simple to operate, low cost, multichannel micro-fluidic chemiluminescence paper chip and the application in detecting at the scene Download PDFInfo
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Abstract
The invention discloses a kind of simple to operate, low cost, the preparation of multichannel micro-fluidic chemiluminescence paper chip and the method for Site Detection thereof.Adopt full printing model, on the ordinary filter paper of an A4 size, bulk print goes out multiple micro-fluidic chemiluminescence paper chip.Print procedure comprises: bulk print hydrophobic wax pattern; Dewaxing is shaping; Bulk print chemical illuminating reagent ink pattern; Bulk print oxidizing ferment ink pattern; The cutting of micro-fluidic chemiluminescence paper chip; The micro-fluidic chemiluminescence paper chip of preparation is carried out plastic packaging process.A method for the Site Detection of micro-fluidic chemiluminescence paper chip, comprises the steps: that the micro-fluidic chemiluminescence paper chip by plastic packaging puts into the magazine of palm luminometer; Sample solution is added drop-wise in sample introduction zone; Then cover cassette cover, start to detect.Judge whether glucose, polyphenol, xanthine, cholesterol, uric acid and ferroheme exist and content successively by the size of 6 the chemiluminescence peak values occurred successively.
Description
Technical field
The present invention relates to low cost, high flux, on-the-spot instant analysis detection technique field simple to operate, is more particularly a kind of with the structure being suitable for the micro-fluidic paper chip laboratory technique platform that chemiluminescence enzyme is analyzed.
Background technology
At present, external (clinical) diagnosis methods mainly contains two kinds: a kind of is the full-automation in the center analysis laboratory utilizing hospital supporting, high-sensitive large-scale instrument and equipment, realizes the diagnosis of high-precision diseases analysis; Another kind of diametrically opposite approach is by the compact simplified equipment of palm, realizes on-the-spot rapid analysis diagnosis.Due to aging population aggravation, the sick incidence of disease of growing up sharply increases, and as relied on merely HC laboratory, greatly will improve national Cost of Health.Therefore, for diagnosing in advance and preventing the generation of major disease, develop that some are simple, low cost, the method and apparatus that can realize family oriented analyzing and diagnosing becomes very urgent.In addition, although people have been devoted to testing cost and the detection time of studying reduction HC analytical test room, when testing result requires higher to ease-to-operate and instantaneity, field quick detection technology is indispensable.
Microfluid based Lab on a chip is the technology platform being expected to realize Site Detection diagnosis at present most.Briefly, Microfluid based Lab on a chip refers to and biological, chemistry, medical analysis process the basic operation unit such as sample preparation, reaction, separation, detection is integrated on the chip of one piece of micro-meter scale, and this chip can be plastic sheet, sheet glass or silicon chip.By building microchannel, micro-valve integrated micro pump technology on these chips, these basic operation units being communicated with to get up, forming Microfluid based Lab on a chip.But at present, Microfluid based Lab on a chip is still in the laboratory research stage, due to its complexity, expensive Preparation equipment and the preparation technology to environmental requirement harshness, the extensive use of Microfluid based Lab on a chip be significantly limit, especially in fund and the underdeveloped developing country of technology.In addition, due to needs, professional person operates, and more cannot realize the analyzing and diagnosing of family oriented.
Paper is a kind of material being dirt cheap, enriching, and test strips is also widely used in the fields such as simple medical diagnosis on disease, environment measuring as a kind of analysis platform (immunochromatography), such as very early pregnancy test strips etc.2007, the Whiteside seminar of Ha Fu university chemistry institute proposes the new ideas in micro-fluidic paper chip laboratory first: refer to biological, chemistry, medical analysis process the basic operation unit such as sample preparation, reaction, separation, detection to be integrated on paper, utilize hydrophobic material on paper, draw out the figure of functional unit and passage, form hydrophilic region (passage).Then, by the capillarity of paper, realize the directed flow of liquid in hydrophilic channel.Micro-fluidic paper chip laboratory has that cost is low, preparation method simply (utilizes simple printing technique, and without the need to harsh conditions such as dust free rooms), (without the need to additional equipment, as pump etc.) simple to operate, use after the advantage such as can to process arbitrarily.
At present, be based upon the analytical method mainly colorimetric method on micro-fluidic paper chip technology platform, because colorimetric method can only provide the signal response of " Yes/No ", and colorimetric method sensitivity is lower, selective poor, easily occurs result false positive.Therefore in micro-fluidic paper chip laboratory, set up the analytical method of high sensitivity, high selectivity, realize high sensitivity, the on-the-spot instant analysis of high specific detects and just become one of this research field problem needing solution badly current.
Summary of the invention
The technical problem to be solved in the present invention is in micro-fluidic paper chip, set up multicomponent chemical Luminescence Enzyme analyzing detecting method that is highly sensitive, high specificity; This micro-fluidic paper chip also has the features such as sample treatment is simple, detection speed is fast, cost is low, and for detecting while glucose, polyphenol, xanthine, cholesterol, uric acid and ferroheme in sample.
In order to solve the problems of the technologies described above, the present invention realizes by building a kind of novel micro-fluidic chemiluminescence paper chip, and the preparation method of this micro-fluidic chemiluminescence paper chip is:
(1) the hydrophobic wax bulk print pattern (pattern as shown in Figure 1) of slight Flow Control chemiluminescence paper chip is designed on computers.
(2) the chemical illuminating reagent bulk print pattern (pattern as shown in Figure 2) with hydrophobic wax bulk print pattern match is designed on computers.
(3) design on computers and six oxidizing ferment bulk print patterns of hydrophobic wax bulk print pattern match (pattern is as shown in accompanying drawing 3,4,5,6,7,8).
(4) filter paper is cut into A4 size.
(5) filter paper cut out in step (4) is placed in wax spray printer, the hydrophobic wax bulk print pattern in step (1) is printed on the filter paper in step (4).
(6) filter paper with wax pattern in step (5) is placed in panel heater or baking oven, under 60-150 DEG C degree Celsius, heats 0.5-2 minute.Wax melted and soaks into the thickness of whole paper, forming hydrophobic wall (principle as shown in Figure 9).
(7) filter paper of preparation in step (6) is put into ink-jet printer.According to the chemical illuminating reagent bulk print pattern in step (2), chemical illuminating reagent ink is printed in chemiluminescence detection district, then according to the oxidizing ferment bulk print pattern in step (3), oxidizing ferment ink is printed in enzyme identification catalytic microchannel.A kind of oxidizing ferment bulk print pattern is used for printing a kind of oxidizing ferment ink: accompanying drawing 3 is for printing glucose oxidase ink; Accompanying drawing 4 is for printing polyphenol oxidase ink; Accompanying drawing 5 is for printing xanthine oxidase ink; Accompanying drawing 6 is for printing cholesterol oxidase ink; Accompanying drawing 7 is for printing urate oxidase ink; Accompanying drawing 8 is for printing heme oxidase ink.When printing different oxidizing ferment ink, only need change the print cartridge in ink-jet printer.
(8) along wax pattern outward flange, cutting is carried out to the filter paper of preparation in step (7), obtain micro-fluidic chemiluminescence paper chip (pattern as shown in Figure 10).
(9) be clipped in the middle of two panels plastic packaging film by obtaining micro-fluidic chemiluminescence paper chip in step (8), then carry out plastic packaging encapsulation, prepare the micro-fluidic chemiluminescence paper chip (plastic packaging stack manner as shown in Figure 11) of encapsulation.
The hydrophilic region (as shown in white portion in accompanying drawing 12) that the hydrophobic pattern (as shown in black part in accompanying drawing 12) of designed micro-fluidic chemiluminescence paper chip is formed comprises a center sample introduction zone (diameter 5.0 ~ 20.0 mm), six in around distribution chemiluminescence detection district (diameter 2.0 ~ 8.0 mm) and six enzyme identification catalytic microchannel (width is 1.0 ~ 4.0 mm, the length of the shortest enzyme identification catalytic microchannel is 4.0 ~ 7.0 mm, other five enzyme identification catalytic microchannel length are 1.5 ~ 6.0 mms longer than previous enzyme identification catalytic microchannel successively.Micro-fluidic chemiluminescence paper chip overall size is 25.0 mm × 25.0, mm ~ 100.0 mm ~ 100.0 mm (square).According to different micro-fluidic chemiluminescence paper chip sizes, the micro-fluidic chemiluminescence paper chip number (7 ~ 30) that an A4 paper prints can be adjusted.
The chemical illuminating reagent print pattern pattern of designed micro-fluidic chemiluminescence paper chip as shown in Figure 13.
Oxidizing ferment print pattern pattern in designed micro-fluidic chemiluminescence paper chip as shown in Figure 14.
Oxidizing ferment print pattern in described accompanying drawing 14 in each enzyme identification catalytic microchannel is of a size of the square of 1.0 ~ 4.0 mm × 1.0 ~ 4.0 mm.
The distance in the oxidizing ferment print pattern distance chemiluminescence detection district in described accompanying drawing 14 in each enzyme identification catalytic microchannel is 2.0 ~ 5.0 mm.
The filter paper adopted is conventional filter paper.
The wax spray printer adopted is conventional Fuji-Xerox's wax spray printer.
The noble metal nano particles solution that chemical illuminating reagent ink described in step (6) is modified for chemical illuminating reagent.The chemical illuminating reagent adopted is conventional chemical illuminating reagent, can be different luminol.The noble metal nano particles adopted is common noble metal nano particles, can be golden nanometer particle.
Oxidizing ferment stamping ink described in step (6) is the noble metal nano particles solution that oxidizing ferment is modified, and the oxidizing ferment adopted is respectively glucose oxidase, polyphenol oxidase, xanthine oxidase, cholesterol oxidase, urate oxidase and heme oxidase.The noble metal nano particles adopted is common noble metal nano particles, can be golden nanometer particle.
Plastic packaging film described in step (8) with sample holes, its size, position identical with paper chip sample introduction zone (pattern is as shown in Figure 11).
The step utilizing the micro-fluidic chemiluminescence paper chip of above-mentioned preparation to realize simultaneously detecting at multi-component scene is:
(1) micro-fluidic chemiluminescence paper chip is put into the magazine of palm luminometer.
(2) sample solution is added drop-wise in the sample introduction zone of micro-fluidic chemiluminescence paper chip.Then cover cassette cover, start to detect.Judge whether glucose, polyphenol, xanthine, cholesterol, uric acid and ferroheme exist and content successively by the size of 6 the chemiluminescence peak values occurred successively.
In the present invention, sample solution under the driving of capillary force, can flow through six enzyme identification catalytic microchannel.Measured object after oxidasic specific recognition catalysis, the hydrogen peroxide of generation flow into chemiluminescence detection district and with chemical illuminating reagent generation chemical reaction, produce optical signal.Because the length of enzyme identification catalytic microchannel is different, cause the time in produced hydrogen peroxide arrival chemiluminescence detection district also different, the luminous signal that different measured object produces can distinguish from the time by this design, furthermore achieved that many groups under a luminous detection window detect simultaneously.
beneficial effect of the present invention:
1. in micro-fluidic paper chip laboratory, introduce highly sensitive chemical luminescence detection method, expanded the detection range in micro-fluidic paper chip laboratory, improve detection sensitivity and the degree of accuracy in micro-fluidic paper chip laboratory.
2. adopt the full preparation mode printed, simplify micro-fluidic chemiluminescence paper chip preparation process, reduce preparation cost, the detection that improve micro-fluidic chemiluminescence paper chip is repeatable.
3. adopted print pattern can carry out large-scale integrated printing, realizes each page printing multiple pattern for preparing multiple micro-fluidic chemiluminescence paper chip simultaneously.
4. contain noble metal nano particles in stamping ink, by the catalytic action of noble metal nano particles, enzyme identification catalytic capability and chemiluminescence efficiency can be improved, improve the sensitivity of this micro-fluidic chemiluminescence paper chip further.
5. construct high-throughout micro-fluidic chemiluminescence paper chip, improve detection flux and the detectability of micro-fluidic chemiluminescence paper chip.
6., due to the porous character of filter paper, therefore enzyme identification catalytic microchannel also has the function that paper chromatography is separated, and is directly filtered out by large granular impurity.The specific recognition catalytic capability of co-oxidation enzyme molecule, this micro-fluidic chemiluminescence paper chip can realize adding detection without the direct sample of pre-treatment, simplifies detecting step, saves sample pre-treatments cost.
7. pair micro-fluidic chemiluminescence paper chip is carried out after plastic packaging, can prevent in transport, store and pollute micro-fluidic chemiluminescence paper chip in use procedure.This design is simultaneously easy to carry and use more.
8. pair micro-fluidic chemiluminescence paper chip carries out after plastic packaging sealing, can improving the REFRIGERATION SYSTEM DRIVEN BY CAPILLARY FORCE power of paper, accelerates the flow velocity of solution in paper passage, reduces detection time further, improve detection efficiency.
Accompanying drawing explanation
Fig. 1. be the hydrophobic wax bulk print pattern of micro-fluidic chemiluminescence paper chip.
Fig. 2. be the chemical illuminating reagent bulk print pattern of micro-fluidic chemiluminescence paper chip.
Fig. 3 is the glucose oxidase bulk print pattern of slight Flow Control chemiluminescence paper chip.
Fig. 4 is the polyphenol oxidase bulk print pattern of micro-fluidic chemiluminescence paper chip.
Fig. 5 is the xanthine oxidase bulk print pattern of micro-fluidic chemiluminescence paper chip.
Fig. 6 is the cholesterol oxidase bulk print pattern of micro-fluidic chemiluminescence paper chip.
Fig. 7 is the urate oxidase bulk print pattern of micro-fluidic chemiluminescence paper chip.
Fig. 8 is the heme oxidase bulk print pattern of micro-fluidic chemiluminescence paper chip.
Fig. 9 is that wax spray prints structure hydrophilic channel principle schematic, and A figure is the schematic diagram of blank filter paper; B figure is the schematic diagram printing wax pattern on filter paper, and wherein a is the wax layer printed; C figure is that in baking oven or flat heater equipment after heating, wax pattern melts and soaks into the thickness of whole filter paper, forms hydrophobic wall.
Figure 10 is the schematic diagram of micro-fluidic chemiluminescence paper chip, and wherein a is the chemiluminescence detection district being printed on chemical illuminating reagent ink, and b is the oxidizing ferment ink printed in enzyme identification catalytic microchannel, and c is sample introduction zone.
Figure 11 is the plastic packaging encapsulation schematic diagram of micro-fluidic chemiluminescence paper chip, and A figure is top view; B figure is sectional view.Wherein a is plastic packaging film, and b is micro-fluidic chemiluminescence paper chip.
Figure 12 is the hydrophobic pattern schematic diagram in micro-fluidic chemiluminescence paper chip, and wherein, 1 is sample introduction zone; 2 is enzyme identification catalytic microchannel; 3 is chemiluminescence detection districts.
Figure 13 is the chemical illuminating reagent print pattern schematic diagram in micro-fluidic chemiluminescence paper chip.
Figure 14 is the oxidizing ferment print pattern schematic diagram in micro-fluidic chemiluminescence paper chip, and wherein, 1 is glucose oxidase ink; 2 is polyphenol oxidase ink; 3 is xanthine oxidase ink; 4 is cholesterol oxidase ink; 5 is urate oxidase ink; 6 is heme oxidase ink.
Detailed description of the invention
Below in conjunction with drawings and Examples, the present invention will be further described.
embodiment 1in human blood, the scene of glucose, polyphenol, xanthine, cholesterol, uric acid and ferroheme is detected fast simultaneously.
(1) the hydrophobic wax bulk print pattern (pattern as shown in Figure 1) of the micro-fluidic chemiluminescence paper chip of Adobe Illustrator CS4 Software for Design is utilized on computers.Size is respectively: center sample introduction zone (diameter 10.0 mm), six in around distribution chemiluminescence detection district (diameter 4.0 mm) and six enzyme identification catalytic microchannel (width is 2.0 mm, and the decibel of six enzyme identification catalytic microchannel is 5.0 mm; 8.0 mm; 11.0 mm; 14.0 mm; 17.0 mm; 20.0 mm.Micro-fluidic chemiluminescence paper chip is of a size of 50.0 mm × 50.0 mm.Now, printable 15 the micro-fluidic chemiluminescence paper chip of A4 paper.
(2) the chemical illuminating reagent bulk print pattern (pattern as shown in Figure 2) of Adobe Illustrator CS4 Software for Design and hydrophobic wax bulk print pattern match is utilized on computers.
(3) six oxidizing ferment bulk print patterns (pattern is as shown in accompanying drawing 3,4,5,6,7,8) of Adobe Illustrator CS4 Software for Design and hydrophobic wax bulk print pattern match are utilized on computers.
(4) common qualitative filter paper is cut into A4 size.
(5) the A4 qualitative filter paper in step (4) is placed in wax spray printer, the hydrophobic wax bulk print pattern in step (1) is printed on the A4 qualitative filter paper cut out in step (4).
(6) the A4 qualitative filter paper with wax pattern in step (5) is placed in panel heater or baking oven, heats 50 seconds under 130 degrees Celsius.
(7) preparative chemistry luminescence reagent ink: get the different luminol aqueous solution of 20.0 milliliters of N-(4-aminobutyl)-N-ethyl (0.5 mole often liter), join (golden nanometer particle diameter is 12.0 nm) in 20.0 milliliters of solution of gold nanoparticles, stirred at ambient temperature 1 hour, rotating speed 500 rpms.Then with the dialysis membrane of 3500 molecular weight and ultra-pure water, reflection solution is dialysed two days, between dialysis period, change six ultra-pure waters.Finally obtain luminol chemiluminescence reagent ink.
(8) prepare oxidizing ferment ink: get 0.5 milliliter of solution of streptavidin (1.0 milligrams every milliliter), join (golden nanometer particle diameter is 12.0 nm) in 20.0 milliliters of solution of gold nanoparticles, stirred at ambient temperature 0.5 hour.Then adding 0.5 milliliter of mass fraction is the bovine serum albumin solution of 5 percent, stirred at ambient temperature 5 minutes.Then by the centrifugation 20 minutes under the rotating speed of 12500 rpms of above-mentioned reaction solution.The precipitation obtained is distributed in biotinylated oxidizing ferment solution, stirs 1 hour under 37 degrees Celsius.By the centrifugation 10 minutes under the rotating speed of 12500 rpms of above-mentioned reaction solution.The precipitation obtained is distributed to (pH=8.0,0.05 mole often liter) in Tris-HCl cushioning liquid, prepares oxidizing ferment ink.
(9) the A4 qualitative filter paper of preparation in step (6) is put into ink-jet printer.First, ink-jet printer is installed chemical illuminating reagent print cartridge, according to the chemical illuminating reagent bulk print pattern in step (2), chemical illuminating reagent ink is printed in chemiluminescence detection district.Then be oxidizing ferment print cartridge by the replacing ink cartridge in ink-jet printer, according to the oxidizing ferment bulk print pattern in step (3), oxidizing ferment ink printed in enzyme identification catalytic microchannel.A kind of oxidizing ferment bulk print pattern is only used for printing a kind of oxidizing ferment ink: accompanying drawing 3 is for printing glucose oxidase ink; Accompanying drawing 4 is for printing polyphenol oxidase ink; Accompanying drawing 5 is for printing xanthine oxidase ink; Accompanying drawing 6 is for printing cholesterol oxidase ink; Accompanying drawing 7 is for printing urate oxidase ink; Accompanying drawing 8 is for printing heme oxidase ink.
(10) carry out cutting with the A4 qualitative filter paper of scissors to preparation in step (9) along wax pattern outward flange, obtain micro-fluidic chemiluminescence paper chip (pattern as shown in Figure 10).
(11) being clipped in the middle of two panels plastic packaging film to obtaining micro-fluidic chemiluminescence paper chip in step (10), then carrying out plastic packaging encapsulation, prepare the micro-fluidic chemiluminescence paper chip (stack manner as shown in Figure 11) of encapsulation.
With the blood sample of three high patients as analysis sample, containing glucose, polyphenol, xanthine, cholesterol, uric acid and ferroheme in blood sample.The step utilizing the micro-fluidic chemiluminescence paper chip of above-mentioned preparation to realize simultaneously detecting at multi-component scene is:
(12) micro-fluidic chemiluminescence paper chip is put into the magazine of palm luminometer.
(13) get patient's blood sample 0.5 milliliter, join (pH=8.0,0.05 mole often liter) in 5 milliliters of Tris-HCl cushioning liquid and dilute.Then with dropper, the sample solution after dilution is added drop-wise in the sample introduction zone of micro-fluidic chemiluminescence paper chip.Then cover cassette cover, start to detect.Judge whether glucose, polyphenol, xanthine, cholesterol, uric acid and ferroheme exist and content successively by the size of 6 the chemiluminescence peak values occurred successively.
embodiment 2the scene of the glucose in human urine, polyphenol, xanthine, cholesterol, uric acid and ferroheme is detected fast simultaneously.
" utilizing Adobe Illustrator CS4 Software for Design " in embodiment 1 step (1) (2) (3) is changed into " utilizing Photoshop CS4 Software for Design "; " qualitative filter paper " in embodiment 1 is changed into " quantitative filter paper "; Embodiment 1 step (1) is changed into: " utilize the hydrophobic wax bulk print pattern (pattern as shown in Figure 1) of the micro-fluidic chemiluminescence paper chip of Photoshop CS4 Software for Design on computers.Size is respectively: center sample introduction zone (diameter 7.5 mm), six in around distribution chemiluminescence detection district (diameter 3.0 mm) and six enzyme identification catalytic microchannel (width is 1.5 mm, and the decibel of six enzyme identification catalytic microchannel is 3.75 mm; 6.0 mm; 8.25 mm; 10.5 mm; 12.75 mm; 15.0 mm.Micro-fluidic chemiluminescence paper chip is of a size of 37.5 mm × 37.5 mm.Now, printable 20 the micro-fluidic chemiluminescence paper chip of A4 paper "." heating 50 seconds under 130 degrees Celsius " in embodiment 1 step (6) is changed into " heating 30 seconds under 150 degrees Celsius "; Embodiment 1 step (7) is changed into " preparative chemistry luminescence reagent ink: get 20.0 milliliters of different luminol aqueous solution (0.5 mole often liter); join (Nano silver grain diameter is 13.0 nm) in 20.0 milliliters of silver nano-particle solution; stirred at ambient temperature 1 hour, rotating speed 500 rpms.Then with the dialysis membrane of 3500 molecular weight and ultra-pure water, reflection solution is dialysed two days, between dialysis period, change six ultra-pure waters.Finally obtain luminol chemiluminescence reagent ink "; " in solution of gold nanoparticles (golden nanometer particle diameter is 12.0 nm) " in embodiment 1 step (8) is changed into " in silver nano-particle solution (Nano silver grain diameter is 15.0 nm) "; " three high patient's blood samples " in embodiment 1 is changed into " three high urine samples "; Embodiment 1 step (12) is changed into and " gets urine sample 5 milliliters, then with common dropper, urine sample is added drop-wise in the sample introduction zone of micro-fluidic chemiluminescence paper chip.Then cover cassette cover, start to detect.Judge whether glucose, polyphenol, xanthine, cholesterol, uric acid and ferroheme exist and content successively by the size of 6 the chemiluminescence peak values occurred successively.”。
Claims (9)
1. a preparation method for multichannel micro-fluidic chemiluminescence paper chip, is characterized in that comprising the following steps:
1.1. the hydrophobic wax bulk print pattern of micro-fluidic chemiluminescence paper chip is designed on computers;
1.2. the chemical illuminating reagent bulk print pattern with hydrophobic wax bulk print pattern match is designed on computers;
1.3. six oxidizing ferment bulk print patterns with hydrophobic wax bulk print pattern match are designed on computers;
1.4. filter paper is cut into A4 size;
1.5. the filter paper cut out in step 1.4 is placed in wax spray printer, the hydrophobic wax bulk print pattern in 1.1 is printed on the A4 filter paper in 1.4;
1.6. the A4 filter paper with wax pattern in 1.5 is placed in panel heater or baking oven, at 60-150 DEG C, heats 0.5-2 minute;
1.7. the A4 filter paper of preparation in 1.6 is put into ink-jet printer; According to the chemical illuminating reagent bulk print pattern in step 1.2, chemical illuminating reagent ink is printed in chemiluminescence detection district, then according to the oxidizing ferment bulk print pattern in step 1.3, utilizing ink-jet printer oxidizing ferment ink to be printed to width is 1.0 ~ 4.0 mm, in the enzyme identification catalytic microchannel that length is different; When printing different oxidizing ferment ink, only need change the print cartridge containing different oxidizing ferment ink in ink-jet printer;
1.8. along wax pattern outward flange, cutting is carried out to the A4 filter paper of preparation in 1.7, obtain micro-fluidic chemiluminescence paper chip;
1.9. being clipped in the middle of two panels plastic packaging film by obtaining micro-fluidic chemiluminescence paper chip in 1.8, then carrying out plastic packaging encapsulation, prepare the micro-fluidic chemiluminescence paper chip of encapsulation.
2. the preparation method of a kind of multichannel micro-fluidic chemiluminescence paper chip according to claim 1, it is characterized in that: the hydrophilic region that the hydrophobic pattern of designed micro-fluidic chemiluminescence paper chip is formed comprises a center sample introduction zone, its diameter is 10.0 mm, six in the chemiluminescence detection district around distribution, its diameter is 2.0 ~ 8.0 mm, six enzyme identification catalytic microchannel, its width is 1.0 ~ 4.0 mm, the length of the shortest enzyme identification catalytic microchannel is 4.0 ~ 7.0 mm, other five enzyme identification catalytic microchannel length are 1.5 ~ 6.0 mms longer than previous enzyme identification catalytic microchannel successively, micro-fluidic chemiluminescence paper chip overall size is the square of 25.0 mm × 25.0, mm ~ 100.0 mm ~ 100.0 mm, according to different micro-fluidic chemiluminescence paper chip sizes, an A4 paper can print 7 ~ 30 micro-fluidic chemiluminescence paper chip.
3. the preparation method of a kind of multichannel micro-fluidic chemiluminescence paper chip according to claim 1, is characterized in that: the diameter of the chemiluminescence pattern in each chemiluminescence detection district is 2.0 ~ 8.0 mm.
4. the preparation method of a kind of multichannel micro-fluidic chemiluminescence paper chip according to claim 1, is characterized in that: the oxidizing ferment pattern in each enzyme identification catalytic microchannel is of a size of the square of 1.0 ~ 4.0 mm × 1.0 ~ 4.0 mm.
5. the preparation method of a kind of multichannel micro-fluidic chemiluminescence paper chip according to claim 1, is characterized in that: the filter paper adopted is conventional filter paper.
6. the preparation method of a kind of multichannel micro-fluidic chemiluminescence paper chip according to claim 1, is characterized in that: the noble metal nano particles solution that described chemical illuminating reagent ink is modified for chemical illuminating reagent; The chemical illuminating reagent adopted is conventional chemical illuminating reagent; The noble metal nano particles adopted is common noble metal nano particles.
7. the preparation method of a kind of multichannel micro-fluidic chemiluminescence paper chip according to claim 1, it is characterized in that: the noble metal nano particles solution that described oxidizing ferment ink is modified for oxidizing ferment, the oxidizing ferment adopted is respectively glucose oxidase, polyphenol oxidase, xanthine oxidase, cholesterol oxidase, urate oxidase and heme oxidase; The noble metal nano particles adopted is common noble metal nano particles.
8. according to the preparation method of a kind of multichannel micro-fluidic chemiluminescence paper chip according to claim 1, it is characterized in that: described two panels plastic packaging film, upper plastic packaging film is wherein with sample holes, and its size, position are identical with paper chip sample introduction zone.
9. application is detected immediately in the scene of micro-fluidic chemiluminescence paper chip that preparation method as claimed in claim 1 obtains, and comprises the following steps:
9.1. micro-fluidic chemiluminescence paper chip is put into the magazine of palm luminometer;
9.2. sample solution is added drop-wise in the sample introduction zone of micro-fluidic chemiluminescence paper chip; Then cover cassette cover, start to detect; Judge whether glucose, polyphenol, xanthine, cholesterol, uric acid and ferroheme exist and content successively by the size of 6 the chemiluminescence peak values occurred successively.
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