CN106596209A - Pretreatment and detection method of dry blood sample - Google Patents
Pretreatment and detection method of dry blood sample Download PDFInfo
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- CN106596209A CN106596209A CN201510661109.4A CN201510661109A CN106596209A CN 106596209 A CN106596209 A CN 106596209A CN 201510661109 A CN201510661109 A CN 201510661109A CN 106596209 A CN106596209 A CN 106596209A
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Abstract
The invention discloses a detection method of a dry blood sample. The detection method comprises the following steps: obtaining a specific blood component-containing dry blood sample to be detected; adding a hemolysis reagent to the dry blood sample to be detected in order to obtain a dry blood sample-to-be-detected and hemolysis reagent mixed solution; and disposing the mixed solution at a specific dissolving temperature for a certain dissolving time in order to obtain a redissolved solution of the dry sample to be detected, wherein the hemolysis reagent comprises a first organic solvent and a second organic solvent, the first organic solvent and the second organic solvent can be different or can be same, and the first organic solvent and the second organic solvent are not methanol when the first organic solvent and the second organic solvent are same. The invention also discloses a method for preparing a standard product of the specific blood component. The invention further discloses a method for detecting the specific blood component in the dry blood sample to be detected.
Description
Technical field
The present invention relates to blood testing field, dry blood sample is pre-processed in particular to one kind,
The method of the specific blood constituent in for detecting dry blood sample and prepare the standard items of specific blood constituent
Method.
Background technology
Blood as human recycle system " logistics " carrier, containing thousands of kinds of protein and small molecule etc. into
Point.For the various composition in blood sample is detected, it will be appreciated that every functional state of human body, this
It is exactly the theoretical foundation of clinical blood detection.And blood clinical detection typically scene extraction at home is fresh
Vein hematometry.
Blood fat is in blood or lipid that is free or being bound to other molecules, generally with triglycerides
(TG), T-CHOL (TC), HDL-C (HDL-C) and low-density lipoprotein
Cholesterol (LDL-C) concentration come characterize individual blood lipid level, be referred to as four items of blood lipid tests.Hyperlipidemia is
The rising or the presence of abnormal level of lipid and/or lipoprotein in blood, it is the main danger of angiocardiopathy
Dangerous factor.
For such as high fat of blood chronic diseases patient, regular monitoring lipids contents are needed.Because scene is taken out
Extracting vein blood is determined has difficult blood sampling, sample transport inconvenience and holding time, and existing
The domestic glucometer degree of accuracy and sensitivity are all often than relatively low, it is impossible to reach the purpose of accurate measurement, therefore compel
Being essential will set up a kind of system of convenient, accurate detection blood sample.
Dried blood spot (DBS) sampling is a kind of easily Blood collection, wherein blood sample by trace in sample
It is in collecting tab and dry.Also, according to Friedewald formula：TC=TG/5+HDL-C+LDL-
C, the concentration level of any one can be calculated according to other three concentration levels for determining in four items of blood lipid tests
Obtain, as long as so measuring wantonly three in TG, TC, HDL-C and LDL-C, it is possible to by application
Formula calculates last.
The content of the invention
This application provides to the standard items done the blood sample method for being pre-processed, prepare specific blood constituent
Method and for detecting dry blood sample in specific blood constituent method, it combines dry blood collection skill
Art, by using the preparation side of the standard items of processing method and specific blood constituent in new dry blood sample
Method, detects to doing the specific blood constituent that includes of blood sample so that collect on a large scale sample,
High precision test and long-term monitoring are possibly realized.
In the one side of the application, there is provided a kind of to doing the method that blood sample is pre-processed, including：
Obtain the dry blood sample comprising specific blood constituent；Hemolyzing reagent is added in the dry blood sample to obtain
State the mixed liquor of dry blood sample and hemolyzing reagent；And the mixed liquor is placed into certain under specific solution temperature
Dissolution time to obtain the redissolution solution of the dry blood sample, the hemolyzing reagent include the first organic solvent with
And second organic solvent, wherein first organic solvent can be different from second organic solvent, or
First organic solvent described in person can be identical with second organic solvent, and when described first organic molten
When agent is identical with second organic solvent, first organic solvent and second organic solvent are not methyl alcohol.
In some embodiments, the specific blood constituent is selected from triglycerides, T-CHOL, high density fat
Protein cholesterol, LDL-C or above-mentioned any combination.
In some embodiments, first organic solvent and the second organic solvent are respectively selected from the following group：Ethanol,
Isopropanol, ether, petroleum ether, n-hexane, acetonitrile or above-mentioned any combination.
In some embodiments, the hemolyzing reagent includes methyl alcohol and acetonitrile.In some embodiments, institute
The content range for stating acetonitrile in hemolyzing reagent is 5%-30% (v/v).In some embodiments, the haemolysis
The content range of methyl alcohol is 70%-95% (v/v) in reagent.In some embodiments, the hemolyzing reagent bag
Include acetonitrile.In some embodiments, the specific blood constituent is triglycerides, T-CHOL or low-density
Lipoprotein cholesterol, the hemolyzing reagent is the mixing of methyl alcohol and acetonitrile, wherein, acetonitrile in the hemolyzing reagent
Content range be the content range of methyl alcohol in 5%-30% (v/v) and the hemolyzing reagent be 70%-95%
(v/v).In some embodiments, the hemolyzing reagent is 100% methyl alcohol.In some embodiments,
The specific blood constituent is HDL-C, and the hemolyzing reagent is 100% acetonitrile.One
In a little embodiments, the hemolyzing reagent further includes PBS.
In some embodiments, the dissolution time is at least 15 minutes to 3 hours.In some embodiment party
In formula, the dissolution time is at least 15 minutes, at least 30 minutes, at least 45 minutes, at least 1 hour,
At least 2 hours or at least 3 hours.In some embodiments, the solution temperature is room temperature or 37 DEG C.
In some embodiments, the solution temperature is room temperature, i.e., 20 ± 5 DEG C.
In some embodiments, the dry blood sample is obtained by the following method：Blood sample quilt
Gather and be stored on dry blood cake piece, and make the blood sample cover the whole dry blood cake piece；And to institute
State dry blood cake piece to be sampled to obtain dry blood sample.In some embodiments, the dry blood cake piece diameter is big
In or equal to 10mm.In some embodiments, the diameter containing blood region of the dry blood cake piece is equal to 10mm.
In some embodiments, the dry blood cake piece is sampled and is less than or equal to obtaining one or more diameters
The dry blood sample of 10mm.In some embodiments, the dry blood cake piece is sampled with obtain one or
The dry blood sample of multiple a diameter of 6mm.In some embodiments, the dry blood cake piece is sampled with
Obtain the dry blood sample of one or more a diameter of 3mm.In some embodiments, the sampling is to pass through
The dry blood cake piece punching is carried out using punching instrument.In some embodiments, the sampling is by right
The region containing blood of the dry blood cake piece carries out what all shearings were carried out.
In some embodiments, the dry blood cake piece is the dry blood scraps of paper.
In some embodiments, the method that the described pair of dry blood sample is pre-processed is further included：Using mixed
Close the specific blood constituent during mixed liquor is fully to dissolve the dry blood sample described in instrument waggle.
In some embodiments, the mixed instrument is rotary mixer, and the rotary mixer is set
It is 10 revs/min -60 revs/min for rotary speed, and rotational time is 5 minutes to 1 hour.One
In a little embodiments, the mixed instrument is oscillator, and the oscillator is arranged to hunting speed for 500
Revs/min -1000 revs/min, and duration of oscillation is -30 minutes 10 minutes.
In other embodiments, the method that the described pair of dry blood sample is pre-processed is further included：Use
Automatic fluid processing platform is equipped with and shakes the mixed liquor to be sufficiently mixed the dry blood sample and haemolysis examination
Agent.In some embodiments, the automatic fluid processing platform includes built-in oscillator, described built-in to shake
Swing device to be arranged to hunting speed is 500 revs/min -1000 revs/min, and duration of oscillation is 10 minutes -30
Minute.
In some embodiments, detect that the mixed liquor determines institute using automation or semi-automatic biochemical instruments
State and whether include in mixed liquor blood constituent to be measured and the content of the blood constituent.
In further aspect of the application, there is provided a kind of method of the standard items for preparing specific blood constituent,
Including：Z random blood sample is obtained, and carries out mixing centrifugation and deposited with isolated upper plasma and lower floor
Thing, wherein Z are more than or equal to 3；Obtain the standard liquid of the described specific blood constituent with known content；With
And mix the deposit with the standard liquid standard items of the specific blood constituent, wherein institute are obtained
It is R to state specific blood component concentration in standard itemss。
In some embodiments, by the deposit with the standard liquid with about 5.8:4.2、5.5:4.5 or
5.2:4.8 ratio mixing is to be obtained the specific blood component concentration as RsStandard items.In some embodiment party
In formula, by the deposit with the standard liquid with 5.5:4.5 ratio mixing is with the prepared specific blood
Constituent concentration is RsStandard items.
In in terms of another of the application, there is provided it is a kind of for the specific blood in dry blood sample to be measured into
Divide the method for being detected, including：Obtain and prepared according to the above-mentioned method pre-processed to dry blood sample to be measured
The redissolution solution of the to be measured dry blood sample for obtaining, wherein the volume of dry blood included in the dry blood sample to be measured
For Vt；Take VtdThe dry blood sample described to be measured of volume is redissolved solution and adds the inspection of the specific blood constituent
Test agent is obtaining sample to be tested solution；The sample to be tested solution is detected to obtain the specific blood
The detected value M of component contentt；And using Calibration equation by the detected value M of the specific blood constituent contentt
It is converted into actual value R of the specific blood constituent contentt。
In some embodiments, it is described for being detected to the specific blood constituent in dry blood sample to be measured
Method further includes the step of obtaining the Calibration equation, including：Obtain N kinds and contain different concentration knowns
Rs={ Rs1、Rs2、Rs3、Rs4…RsNSpecific blood constituent the dry blood sample of standard items, wherein N >=5,
And the volume of included dry blood is V in the dry blood sample of the standard itemss={ Vs1、Vs2、Vs3、Vs4…VsN}；
Prepare the redissolution solution of the dry blood sample of N kinds standard items；Take VsdThe dry blood sample of the N kinds standard items of volume
Redissolution solution, and being separately added into the detection reagent of the specific blood constituent, to obtain N kind standard items samples molten
Liquid；The N kinds standard items sample solution is carried out to detect the detected value for obtaining wherein specific blood constituent content
Ms={ Ms1、Ms2、Ms3、Ms4…MsN}；And according to concentration known RsWith detected value MsBetween relation
Obtain normal equation.
In some embodiments, the utilization Calibration equation is by the detected value of the specific blood constituent content
MtIt is converted into actual value R of the specific blood constituent contenttProcess include：By MtInstitute is substituted into as y values
In stating Calibration equation, calculate and obtain x values；And obtain the specific blood by the way that x values are carried out into volume correction
Actual value R of component contentt。
In some embodiments, the Vtd=Vsd, by the way that the x values are multiplied by into Vt/VsTo carry out the body
Product correction.
In some embodiments, the VtdIt is not equal to Vsd, by the way that the x values are multiplied by into (Vt/Vs)*(Vsd/Vtd)
To carry out the volume correction.
In some embodiments, it is described for being detected to the specific blood constituent in dry blood sample to be measured
Method further includes to prepare containing concentration known RsDescribed specific blood constituent standard items dry blood samples
Step, including：Z random blood sample is obtained, and carries out mixing and be centrifuged with isolated upper plasma with
Surface sediments, wherein Z are more than or equal to 3；The standard for obtaining the described specific blood constituent with known content is molten
Liquid；And mix the deposit with the standard liquid so that the standard items of the specific blood constituent are obtained,
Specific blood component concentration is R in wherein described standard itemss。
In some embodiments, the deposit is mixed with 5.5 with the standard liquid:4.5 ratio with
It is R that the specific blood component concentration is obtainedsStandard items.
In some embodiments, after the standard items of the specific blood constituent are obtained, further include：
By the standard items point sample of the specific blood constituent and it is stored in dry blood cake piece, and it is whole to cover the blood sample
The individual dry blood cake piece；And the dry blood cake piece is sampled a including V to obtainbThe dry blood of volume
The dry blood sample of the standard items of amount.In some embodiments, the dry blood sample to be measured is by with lower section
What method was obtained：Blood sample is collected and is stored on dry blood cake piece, and makes the blood sample uniform fold whole
The individual dry blood cake piece；And the dry blood cake piece is sampled a including V to obtainsThe dry blood of volume
The dry blood sample described to be measured of amount.
In some embodiments, the specific blood constituent is selected from the group：T-CHOL, triglycerides, height
Density lipoprotein-cholesterol, LDL-C or above-mentioned any combination.
The method of the pretreatment of the above-mentioned dry blood sample that the present invention is provided, it is possible to use specific hemolyzing reagent
(mixed solvent of such as methyl alcohol and acetonitrile, or pure acetonitrile etc.) quickly and efficiently extract the spy done in blood sample
Determine blood constituent (such as T-CHOL, triglycerides, HDL-C, low-density lipoprotein
Cholesterol or its any combination) content so that using dry blood sample specific blood constituent is carried out in a large number,
Efficiently, accurately detection is possibly realized.
Further, present invention discover that：By adjusting solution temperature, dissolution time, and can be by using
Mixed instrument and/or oscillator, make dry blood sample instinct farthest be dissolved in hemolyzing reagent, so that i.e.
The content of specific blood constituent just can be exactly measured using a small amount of dry blood sample, whole experiment is improve
The degree of accuracy, and cause small sample amount detection be possibly realized.
In addition, the method for detecting the specific blood constituent in dry blood sample is provided in the present invention, its knot
Close dry blood acquisition technique, by using processing method in new dry blood sample to do blood sample include it is specific
Blood constituent detected, so that collect that sample, high precision test and long-term monitoring become on a large scale can
Energy.
Finally, present invention also offers the method for preparing standard items for specific blood constituent, so as to for upper
The detection of the specific blood constituent in stating to doing blood sample provides more accurately calibration reference, so as to improve
The degree of accuracy of whole experiment.
It is above the general introduction of the application, may there is the situation of simplified, summary and omissions of detail, therefore this area
Technical staff it should be appreciated that the part is only Illustrative, and is not intended to limit by any way
The application scope.This overview section is both not intended to determine the key feature of claimed subject or necessary special
Levy, the supplementary means of the scope for being also not intended to be used as to determine claimed subject.
Description of the drawings
Combined by following description and appending claims and with accompanying drawing, it will be described more fully
The above and other feature of teachings herein.If it is appreciated that these accompanying drawings depict only teachings herein
Dry embodiment, therefore it is not considered as the restriction to teachings herein scope.By adopting accompanying drawing, this Shen
Please content will obtain definitely and detailed description.
Fig. 1 shows a kind of schematic diagram of the applicator of specific embodiment of the invention.
Fig. 2 shows a kind of method of the pretreatment of the dry blood sample of specific embodiment of the invention
Exemplary process diagram.
Fig. 3 shows that a kind of preparing for specific blood constituent for specific embodiment of the invention is marked
The exemplary process diagram of the method for quasi- product.
Fig. 4 show a kind of specific embodiment of the invention for detecting dry blood sample to be measured in
The exemplary process diagram of the method for specific blood constituent.
Fig. 5 shows a kind of example of the method for the acquisition Calibration equation of specific embodiment of the invention
Property flow chart.
Fig. 6 show another kind of specific embodiment of the invention for detecting dry blood sample to be measured in
Specific blood constituent method exemplary process diagram.
Fig. 7 A-7C are respectively illustrated when the mixed solution of the acetonitrile of 75% methyl alcohol+25% is used as hemolyzing reagent,
The correlation of the corresponding WBS detected value of the dry blood examination measured value of TC, TG and LDL-C.
Fig. 8 A-8C are respectively illustrated when 100% methyl alcohol is used as hemolyzing reagent, TC, TG and LDL-C's
The correlation of the corresponding WBS detected value of dry blood examination measured value.
When Fig. 9 A-9C are respectively illustrated using hole knockout, the dry blood examination measured value of TC, TG and LDL-C with
The correlation of its corresponding WBS detected value.
When Figure 10 A-10C are shown using cut mode, the dry blood examination measured value of TC, TG and LDL-C and its
The correlation of corresponding WBS detected value.
Specific embodiment
In the following detailed description, with reference to constitute part thereof of accompanying drawing.In the accompanying drawings, similar symbol
Number similar part is generally represented, unless otherwise indicated by context.Describe in detail, accompanying drawing and right will
The illustrative embodiments described in book are asked to be not intended to limit.The theme without departing from the application spirit or
In the case of scope, other embodiment can be adopted, and other changes can be made.It is appreciated that
General various aspects describe, teachings herein illustrating in the accompanying drawings herein can be carried out
Various differently composed configurations, replacement, combination, design, and it is all these all clearly constitute the application in
A part for appearance.
Hereinafter, in order to clearly be described to specific embodiment, will be using some specific terms.So
And using the protection domain for being not intended that restriction the application of these terms, the scope of these terms should be extended to
Any equivalent that roughly the same purpose is reached with roughly the same means.
Dry blood cake piece
For crowd to be taken a blood sample, by using the applicator including dry heme, such as dry blood cake
Piece (including the dry blood scraps of paper and dry blood polymer plastic tablet, dry blood glass and mixed materials carrier etc.), the dry sea of blood
Continuous and dry blood cotton etc., facilitates crowd oneself to take a blood sample, without the need for medicine blood sampling, saving time and efforts, and
And a small amount of finger tip blood volume is only needed to, also allow for collecting great amount of samples.And blood sample is the fortune of dry blood sample
It is defeated also very convenient, it is that patient and laboratory save cost.In some embodiments, the applicator
For dry blood cake piece.In some embodiments, the dry blood cake piece is the dry blood scraps of paper.
Fig. 1 is to be illustrated a kind of specific embodiment of the invention with applicator as dry blood cake piece
Dry blood cake piece 100 schematic diagram.
As shown in figure 1, the dry blood cake piece 100 includes blood sample collection area 101 and sample information area 102.Its
Middle blood sample collection area 101 is provided with the slide glass (for example, filter paper) for collection blood sample, the sample information area 102
Be provided with for fill in sample information corresponding with the blood sample (for example provide the name of the patient of the sample, cell-phone number,
Address and date collected etc.).Although as shown in figure 1, blood sample collection area 101 includes the confession collection of 3 circles
The filter paper of blood sample, but those skilled in the art can according to actual needs arrange number, the shape of filter paper
And size.
Preferably, the dry blood cake piece 100 also includes protection portion 103 and connection side 104.The wherein protection portion
1103 have the dry blood cake piece after cleaning, the protection sampling for keeping blank blood sample collection area 101 not contaminated
Effect；And protection portion 103 connects blood sample collection area 101 by connection side 104, forms integral structure,
The protection portion 103 bends relative to connection side 104, and the blank blood sample collection area 101 is covered.It is logical
Cross the mode for folding to be covered in blood sample collection area 101, to blank sampling face or will not gather what is obtained after blood sample
Dry blood cake piece surface produces friction, and then affects testing result.
In some embodiments, blood sample collection area 101 includes multiple circular Waterman for collection blood sample
Filter paper, self-disinfecting and gathers blood sample (such as finger tip blood), by blood sample drops at home by patient
Enter on the filter paper in blood sample collection area 101 and make it at least to cover the area of whole filter papers, it is further natural
Dry 1-2 hours.In some embodiments, the saturated capacity of blood is adsorbed in the filter paper unit area
It is fixed.In some embodiments the circular Waterman filter papers for collection blood sample are a diameter of
12.5mm, but because neighboring area is unsuitable for storing up blood, the region containing blood of the dry blood cake piece is about diameter
10mm.And thereon the saturated capacity of adsorbable blood is about 40-45 μ l.In some embodiments, one
In the fixed time cycle, regular, multi collect blood, and cause the blood sample difference nature of collection every time full
With absorption on a series of dry blood cake piece, and it is further dried.Also, every time will within 2 days after blood sampling
Dry blood cake piece is delivered to corresponding testing agency by modes such as express deliveries.
Dry blood sample preprocess method
Fig. 2 shows a kind of method 200 of the pretreatment of the dry blood sample of specific embodiment of the invention
Exemplary process diagram.Methods described 200 is comprised the following steps：Obtain the dry blood sample comprising specific blood constituent
This (201)；Hemolyzing reagent is added in the dry blood sample to obtain the dry blood sample and hemolyzing reagent
Mixed liquor (202)；And place certain dissolution time under specific solution temperature to obtain by the mixed liquor
The redissolution solution (203) of the dry blood sample.
The term " specific blood constituent " for using in this application refers to triglycerides, T-CHOL, high density
Lipoprotein cholesterol, LDL-C or its any combination.
" hemolyzing reagent " being related in the application includes being respectively selected from the following group：Ethanol, isopropanol, ether, stone
First organic solvent and the second organic solvent of oily ether, n-hexane, acetonitrile or above-mentioned any combination.At some
In embodiment, first organic solvent is different from second organic solvent.In some embodiments
In, first organic solvent can be identical with second organic solvent, and now described first has
Machine solvent and second organic solvent are not methyl alcohol.In some embodiments, the hemolyzing reagent includes first
Alcohol and acetonitrile.In some embodiments, the content range of acetonitrile is 5%-30% (v/v) in the hemolyzing reagent.
In some embodiments, the content range of methyl alcohol is 70%-95% (v/v) in the hemolyzing reagent.At some
In embodiment, the hemolyzing reagent is that the content range of acetonitrile in methyl alcohol and acetonitrile, and the hemolyzing reagent is
5%-30% (v/v), the content range of methyl alcohol is 70%-95% (v/v) in the hemolyzing reagent.In some enforcements
In mode, the specific blood constituent be triglycerides, T-CHOL or LDL-C, it is described
Hemolyzing reagent is methyl alcohol and acetonitrile, wherein, the content range of acetonitrile is 5%-30% (v/v) in the hemolyzing reagent
And the content range of methyl alcohol is 70%-95% (v/v) in the hemolyzing reagent.In some embodiments, institute
Hemolyzing reagent is stated including acetonitrile.In some embodiments, the hemolyzing reagent is pure methyl alcohol.In some enforcements
In mode, the specific blood constituent is HDL-C, and the hemolyzing reagent is 100% second
Nitrile.In some embodiments, the hemolyzing reagent further includes PBS.Above-mentioned specific hemolyzing reagent can
So that effectively the specific blood constituent in blood sample is dissolved in hemolyzing reagent.
" dry blood sample " described herein refers to that one or more are loaded with dry blood by what is obtained on dry blood cake piece
The sample of blood sample.In some embodiments, described " dry blood sample " can by the following method be obtained：
Blood sample is collected and is stored in the blood sample collection area 101 of dry blood cake piece 100 as above, and makes institute
The area that blood sample at least covers all blood samples pickup area (i.e. region containing blood) in blood sample collection area 101 is stated,
And by such as whole sample collection areas etc. being punched or being cut out to the dry blood cake piece just using punching instrument
Method the dry blood cake piece is sampled with obtain one or more (such as 1,2,3,4,5
It is individual, 6,7,8,9 or more) the dry blood sample, wherein present document relates to dry blood sample can
Be dry blood sample to be measured can also be the dry blood sample of standard items.
In some embodiments, the dry blood sample be diameter less than or equal to 10mm (such as 10mm,
8mm, 7mm, 6mm, 5mm, 4mm, 3mm, 2mm, 1mm) the filter paper for being loaded with dry blood blood sample.
The diameter of the dry blood sample of circle that those skilled in the art are also contemplated that simultaneously is less, wherein included is dry
The volume of blood is less, it is thus possible to which need to obtain the dry blood sample of the circle of more is carried out enough with ensureing sample size
The detection of follow-up specific blood constituent.In some embodiments, the dry blood of a diameter of 10mm of said one
The volume of included dry blood is to be not less than 25 μ l in sample.
It should be understood that total dry blood volume that one or more dry blood samples include (can correspond to always sample as described above
Area) and hemolyzing reagent volume ratio can affect the dissolution rate of the specific blood constituent, dissolution efficiency with
And concentration of the specific blood constituent in dissolution fluid.So in following experimental procedure, be required to ensure for
When the dry blood sample of dry blood sample to be measured and standard items is detected, the gross area of the dry blood sample for being taken, added
Hemolyzing reagent and detection reagent content it is consistent.
In step 202., hemolyzing reagent is added in dry blood sample with reagent adding set described to obtain
The mixed liquor of dry blood sample and hemolyzing reagent.The reagent adding set can be at pipettor or automatic fluid
Platform.In some embodiments, pipettor is manual pipettor.In some embodiments, it is manual to move
Liquid device can be single track pipettor or multichannel pipettor.In some embodiments, using can in batches by haemolysis
Reagent adds the automatic fluid processing platform (such as Agilent Bravo 96LT) in dry blood sample dry to obtain
The mixed liquor of blood sample and hemolyzing reagent.
In step 203, the mixed liquor of the dry blood sample and hemolyzing reagent places one under specific solution temperature
Fixed dissolution time is obtaining the redissolution solution of the dry blood sample.It will be appreciated by those skilled in the art that molten
After blood reagent is added in dry blood sample, spy can be adjusted by adjusting solution temperature and/or dissolution time
Determine efficiency and content that blood constituent is dissolved in hemolyzing reagent, can also be mixing by way of manually or mechanically
Hemolyzing reagent and dry blood sample, to improve efficiency and content that specific blood constituent is dissolved in hemolyzing reagent.One
In a little embodiments, the dissolution time is at least 15 minutes to 3 hours.In some embodiments, it is molten
The solution time is at least 15 minutes, at least 30 minutes, at least 45 minutes, at least 1 hour, at least 2 hours
Or at least 3 hours.In some embodiments, solution temperature is room temperature or 37 DEG C.In some embodiments
In, solution temperature is room temperature, i.e., 20 ± 5 DEG C.Some preferred embodiment in, solution temperature is room temperature,
Dissolution time is 2 hours.
In some embodiments, step 203 still further comprise using the dry blood sample of mixed instrument waggle and
The mixed liquor of hemolyzing reagent fully to dissolve the dry blood sample in specific blood constituent.Wherein it is possible to pass through
The speed of mixed instrument is set to improve efficiency and content that specific blood constituent is dissolved in hemolyzing reagent.In some realities
In applying mode, the mixed instrument is rotary mixer (such as DHS JC502-DR-MIX) or oscillator.
In some embodiments, the rotary mixer can be arranged to rotary speed at 10 revs/min -60 revs/min
In the range of, rotational time is in the range of 5 minutes to 1 hour.In some embodiments, in step 202.
Hemolyzing reagent is added in dry blood sample using automatic fluid processing platform, and the automatic fluid processing platform
Can be used to be sufficiently mixed dry blood sample and hemolyzing reagent in step 203 with built-in oscillator, to improve spy
Determine efficiency and content that blood constituent is dissolved in hemolyzing reagent.In some embodiments, above-mentioned oscillator can quilt
Hunting speed is set in the range of 500 revs/min -1000 revs/min, duration of oscillation 10 minutes -30 points
In the range of clock.
Standard items preparation method
Fig. 3 shows that a kind of preparing for specific blood constituent for specific embodiment of the invention is marked
The schematic diagram of the method 300 of quasi- product.Shown in the calibration steps of following article, present invention also offers one kind is right
Dry blood sample to be measured has carried out pre-processing and obtain after the redissolution solution of dry blood sample to be measured, to redissolving solution
Detected so as to obtain measured value, and for method that the measured value is calibrated using Calibration equation.
Wherein, in order to determine Calibration equation, the R for containing different concentration knowns using N kinds is neededs={ Rs1、Rs2、
Rs3、Rs4…RsNSpecific blood constituent standard items sample.The following describes the side for preparing standard items sample
Method.
The method 300 for preparing standard items for specific blood constituent is comprised the following steps：Obtain Z
Random blood sample, and mixing centrifugation is carried out with isolated upper plasma and seabed sediment (301)；Obtain
Take the standard liquid (302) of the described specific blood constituent with known content；By the deposit with it is described
Standard liquid is mixed to prepare specific blood component concentration as R with certain proportionsStandard items (303).
The blood sample that the random blood sample obtained wherein in step 301 is processed for Jing anti-freezings, random sample
Amount Z be not less than 3, carry out mixing the seabed sediment that obtains of centrifugation be mainly red blood cell additionally include it is white
Cell and blood platelet etc..Generally, plasma content is 55 ± 3% (v/v) in blood of human body, and lower floor sinks
Product thing content is 45 ± 3% (v/v).The content ratio of blood plasma and seabed sediment slightly has in the blood of different people
Difference, but when sample size larger (such as >=100, >=500, >=1000, >=2000 etc.), by each blood sample
The ratio that the upper plasma and seabed sediment for obtaining is centrifuged after this mixing is about 5.5:4.5.
" content " being related in the application includes mass percent, percent by volume, number percent, matter
Amount, molal quantity, mass concentration or molar concentration etc..
In step 302, the commercially available described specific blood constituent with known content of purchase can be passed through
Standard liquid (artificial serum of TG, TC, LDL-C, HDL-C of such as concentration known) is utilized
The dry powder of the described specific blood constituent of known quality or known molal quantity or the concentrate of concentration known etc. are configured
Obtain the standard liquid of the described specific blood constituent with known content.
In step 303, by the deposit with the standard liquid with certain proportion (such as with about
5.8:4.2、5.5:4.5 or 5.2:4.8 ratio) specific blood component concentration is mixed to prepare for RsStandard items.
Skilled artisan would appreciate that concentration R of specific blood constituent hereinsWith the mark for obtaining in step 302
" known content " of quasi- solution is simultaneously differed, and it is poor by ratio caused by mixed proportion to exist between the two
It is different, for example set in step 302 obtain specific blood constituent TC standard liquid in known TC contents as
5mmol/L, the standard liquid obtained in the deposit and the above-mentioned steps 302 that will obtain in step 301 with
5.5:4.5 standard items for being mixed to prepare TC, now in the standard items TC concentration (mmol/ μ l) RsFor
2.25mmol/L.It should be appreciated that specific blood component concentration RsUnit can for mmol/L, mg/dl,
The arbitrarily concentration unit well known in the art such as pmol/ μ l.
In some embodiments, the above-mentioned method 300 for preparing standard items can further include and will make
Standard items point sample, be stored on dry blood cake piece, and to it after standard items on dry blood cake piece are dried
It is sampled, (is specifically referred to above to method 200 so as to obtain the step of standard items do blood sample
The relevant portion of the preparation method of " dry blood sample " in description).
The method of specific blood constituent in the dry blood sample of detection
Fig. 4 show a kind of specific embodiment of the invention for detecting dry blood sample to be measured in
The schematic diagram of the method 400 of specific blood constituent.It is described for detecting dry blood sample to be measured in specific blood
The method 400 of composition includes step：Obtain according to prepared by the method for the pretreatment to dry blood sample to be measured treating
Survey the redissolution solution (401) of dry blood sample；Take VtdThe redissolution solution of the dry blood sample described to be measured of volume is simultaneously
The detection reagent of the specific blood constituent is added to obtain sample to be tested solution (402)；Test sample is treated to described
This solution is detected to obtain the detected value M of the specific blood constituent contentt(403)；Using calibration
Equation is by detected value MtIt is converted into actual value R of specific blood constituent content in sample to be testedt(404)。
In step 401, the step of acquisition can be included by such as the concrete steps described in method 200
Only include directly to prepare the preparation process of the redissolution solution of dry blood sample to be measured, or not include preparing
The behavior (such as purchase is obtained, pays acquisition etc.) of acquisition.
In step 402, V is takentdThe redissolution solution of the dry blood sample described to be measured of volume simultaneously adds the spy
The detection reagent of blood constituent is determined to obtain sample to be tested solution.It will be appreciated by those skilled in the art that can be with
Estimated by experience, mathematics is calculated or experiment test determines taken volume V for redissolving solutiontd, so that
Obtain and redissolving the final concentration of specific blood constituent in the mixed liquor of solution and detection reagent in the concentration for being adapted to detection
In the range of, the concentration range of the suitable detection is received to include instrument detection sensitivity, reagent detection range, dry blood
Sample sampling amount is (such as dry blood volume V for wherein includingt/ dry blood sample gross area St) etc. any art technology
The restriction of factor known in personnel." detection reagent " that the application is related to refer to it is any of or commercially available can
For the reagent of the detection of specific blood constituent.
In step 403, the sample to be tested solution is detected and is contained with obtaining the specific blood constituent
The detected value M of amountt, wherein " detected value " refers to the letter obtained by apparatus measures in medical test
Number in the measure of specific blood constituent content obtained after certain built-in data processing or program conversion
Value.
In step 404, using Calibration equation by detected value MtIt is converted into specific blood constituent in sample to be tested
Actual value R of contenttThe step of completed by mathematical transformation, wherein Calibration equation can pass through art technology
Any mode is obtained known to personnel, and such as Calibration equation can be by using the standard items of concentration known same
Measured value in machine and same detection batch is obtained with the relation of concentration known.
Fig. 5 shows a kind of method 500 of the acquisition Calibration equation of specific embodiment of the invention
Schematic diagram.The method 500 for obtaining Calibration equation includes step：Obtain N kinds and contain different concentration knowns
RsSpecific blood constituent the dry blood sample (501) of standard items；Prepare answering for the dry blood sample of N kinds standard items
Solution (502)；Take VsdThe redissolution solution of the dry blood sample of the N kinds standard items of volume, and add respectively
The detection reagent for entering the specific blood constituent obtains N kinds standard items sample solution (503)；To the N
Planting standard items sample solution carries out detecting the detected value M for obtaining wherein specific blood constituent contents={ Ms1、
Ms2、Ms3、Ms4…MsN}(504)；According to concentration known RsWith detected value MsBetween Relation acquisition
Normal equation (505).
In step 501, the specific blood constituent for containing in the dry blood sample of N kinds various criterion product it is known dense
Degree Rs={ Rs1、Rs2、Rs3、Rs4…RsN, in the dry blood sample in wherein N >=5, and the N kinds standard items
Volume V of included dry bloods={ Vs1、Vs2、Vs3、Vs4…VsN}。
The step of redissolution solution that the dry blood sample of N kinds standard items is prepared in step 502, can refer to dry blood sample above
Specific descriptions in this preprocess method 200.
Step 503 in method 500,504 corresponding to the step 402 in method 400,403；Concrete behaviour
Make details referring to the description in method 400 to corresponding step.
In step 505, according to concentration known RsWith detected value MsBetween Relation acquisition normal equation
Process is completed by data processing, specifically with the Rs={ Rs1、Rs2、Rs3、Rs4…RsNIn it is each
It is individual for abscissa and the Ms={ Ms1、Ms2、Ms3、Ms4…MsNIn corresponding one be ordinate obtain
N number of calibration point, and according to the distribution situation of above-mentioned N number of calibration point, it is determined that reflection concentration known RsAnd inspection
Measured value MsBetween mapping relations Calibration equation.Above-mentioned Calibration equation can be it is any type of, for example linearly
Equation, exponential equation, logarithmic equation etc..In some embodiments, Calibration equation is linear, and should
Linear equation is in x, y-axis without intercept.In some embodiments, Calibration equation is linear, and should
Linear equation intercept on the y axis be positive number (i.e. specific blood constituent actual content be 0 when, due to
The detection background values that the reason for detection method or detection reagent produces).
Fig. 6 show another kind of specific embodiment of the invention for detecting dry blood sample to be measured in
Specific blood constituent method 600 schematic diagram.Step 601-604 wherein in method 600 is corresponded to
Step 401-404 in method 400, concrete operations details to corresponding step in method 400 referring to retouching
State；Method 600 further includes the step of obtaining Calibration equation (603 '), and it corresponds to method 500
In step 501-505, concrete operations details is referring to the description in method 500 to corresponding step.At some
In embodiment, method 600 still further comprises preparation N kinds and contains different concentration knowns RsSpecific blood
The step of standard items of composition (601 '), it corresponds to step 301-303 in method 300, concrete operations
Details is referring to the description in method 300 to corresponding step；Contain different concentration knowns R with by above-mentioned N kindss
The standard items of specific blood constituent be prepared into N kinds and contain different concentration knowns RsSpecific blood constituent mark
The step of dry blood sample of quasi- product (602 '), its concrete operation step can refer to described in method 200
The preparation method of " dry blood sample ".
Hereinafter the present invention will be specifically described by experimental example.
Experimental example
The present invention is further described by following non-limitative experiment examples.It should be noted that enumerating these experimental examples
It is only intended to further illustrate the technical characteristic of the present invention, it is not intended that can not be interpreted to the present invention's
Limit.The experimental example is not comprising the conventional method (chemical synthesising technology well known to persons skilled in the art
Deng) detailed description.
The instrument that the present invention is used is the full-automatic biochemicals of Amorsino 460 point of Aino Electronic Co., Ltd., Shanghai
Analyzer.
In existing clinical labororatory, for the standard method that the detection of high fat of blood is used is based on whole blood
The four items of blood lipid tests detection of (such as fresh venous blood).Dry blood is based primarily upon in this application and uses Shanghai
The four items of blood lipid tests that the automatic clinical chemistry analyzers of Amorsino 460 of Ai Nuo Electronics Co., Ltd.s are determined, and one
Divide in control experiment is to determine four items of blood lipid tests based on WBS.
Embodiment 1：General experimental implementation
The collection and preparation of sample
The preparation of dry blood sample to be measured：Experimenter's finger is carried out disinfection, and is pushed by pin puncture finger tip of taking a blood sample
Finger root blood vessel, makes finger tip blood flow out naturally, and is added dropwise on dry blood cake piece the filter paper for collecting blood sample
In region, the drop of blood amount that wherein finger tip flows out at least should can infiltrate and cover the area of whole filter papers, afterwards
Further about 1-2 hour of drying at room temperature is until the filter paper of infiltration blood is parched.Afterwards using low-permeable
Plastic cement sample sack seals dried dry blood cake piece up for safekeeping, and alternatively in sample sack includes drier to guarantee sample
This keeps drying in storage transportation.Blood-borne pathogens, institute may be included in due to being loaded with blood sample
Answer handled, where in office all should keep using gloves when managing dry blood cake piece with whole experiment process.
Before detection the dry blood cake piece is punched using punching instrument, or go out whole filter paper sections using scissor cut
Domain is sampled to obtain dry blood sample of one or more diameters between 1mm-10mm to the dry blood cake piece
This.
Blood-sampling method of the whole blood blood as control：For each experimenter gathers fresh venous blood sample,
Its blood sample is divided into two parts, first part is whole blood blood, is stored in the heparin tube containing anti-coagulants and is placed in
4 DEG C of refrigerations are standby, for determining the concentration based on the specific blood constituent of whole blood sample；Second part is then used to treat
The preparation of dry blood sample is surveyed, that is, is pipetted and is instilled in the filter paper panel region for being used for collecting blood sample on dry blood cake piece, made
Blood sample infiltrates and covers the entire area (such as the border circular areas of about a diameter of 10mm) of filter paper, afterwards
About 1-2 hour of drying at room temperature, obtains being loaded with the patch of dried blood sample, for determining the spy based on dry blood sample
Determine the concentration of blood constituent.
The pretreatment (dry blood sample to be measured is identical with the preprocess method of the dry blood sample of standard items) of dry blood sample
Dry blood sample is placed in the microcentrifugal tube of 1.5ml and is added 750 μ l hemolyzing reagents.Will be above-mentioned micro
Centrifuge tube is placed on rotary mixer DHS JC502-DR-MIX, and rotary speed is set to into 30 revs/min
Clock, rotational time is set to 1 hour, and a hour is dissolved under room temperature condition, obtains and redissolves solution.
The detection of specific blood constituent
The detection reagent adopted in the application is mainly derived from Li Deman (Leadman), and (T-CHOL is determined
Kit-TC, article No.：TC7150；Triglyceride determination kit-TG, article No.：TG7160；It is highly dense
Degree lipoprotein cholesterol determines kit-HDL-C, article No.：HD7173；Low-density LP determination reagent
Box-LDL-C, article No.：) and the last nine (BSBE) (T-CHOL detection kit, article No. LD7182：
GS101Z；Triglycerides detection kit, article No.：GS111Z；HDL detection kit,
Article No.：GS131Z；Low-density lipoprotein detection kit, article No.：GS141Z), wherein special to each
The testing process of blood constituent is determined, except sample process and consumption part need to be adjusted according to different sample types
Beyond whole, other are carried out according to the operation manual of each detection kit and Amorsino 460 substantially.
In the detection of the specific blood constituent of dry blood sample, according to the operation manual of each detection kit
It is interior to use sample amount for Virus monitory, the consumption (example for redissolving solution is suitably adjusted with its 4-5 times consumption
Such as, if being the μ l detection reagents of 1 μ l serum+199 for serum, can redissolve for 5 μ l for redissolving solution
The μ l detection reagents of solution+195), detected using the automatic clinical chemistry analyzers of Amorsino 460 afterwards.
Embodiment 2：The selection of hemolyzing reagent
Collection and preparation according to the sample described in embodiment 1, the pretreatment of dry blood sample and to specific blood
The method that liquid composition is detected, wherein the total people of experimenter 10 (being represented with S1-S10), respectively takes whole blood
Sample is a and multiple dry blood cake pieces, the disk of some a diameter of 10mm is obtained by shearing, per 3 circles
Piece add a 1.5ml microcentrifugal tube in and add 750 μ l hemolyzing reagents to be dissolved.Wherein use
Hemolyzing reagent is：
First group：PBS is plus in methyl alcohol, ethanol, isopropanol, ether, petroleum ether, n-hexane, acetonitrile
It is arbitrary, wherein both ratios include 1:9、2:8、5:5, totally 21 kinds；
Second group：Methyl alcohol is plus appointing in ethanol, isopropanol, ether, petroleum ether, n-hexane, acetonitrile
One, wherein both ratios include 8:2、6:4、4:6、2:8, totally 24 kinds.
Above-mentioned each group hemolyzing reagent be used to dissolve the specific blood constituent in dry blood sample, and its dissolved efficiency leads to
Cross that the dissolving of more dry blood sample obtains redissolve in solution with WBS in specific blood constituent detected value
To calculate.Distinguished using TC7150, TG7160, HD7173, LD7182 kit of above-mentioned Li Deman
Detect the specific blood constituent dissolved efficiency of above-mentioned first group and second group hemolyzing reagent.Wherein first group PBS
There is certain effect in fraction with the mixed system of other organic reagents, but be not optimal.At second group
Methyl alcohol with the mixed system of other organic reagents in, methyl alcohol and hexane, methyl alcohol and ethanol, methyl alcohol and second
The part mixed proportion of nitrile, isopropanol and methyl alcohol is for some blood constituents (such as TC, TG, LDL-
C) there is relatively high dissolved efficiency (data do not include).But with regard to utilizing in conceptual data and prior art
100% methyl alcohol is carried out for the result that the efficiency of haemolysis is compared, wherein methyl alcohol (MeOH) and acetonitrile
(ACN) mixed system compares other significantly more dissolved efficiency compared with the system of 100% methyl alcohol
Raising.The inspection of another group of solubilising reagent is carried out with the optimum proportioning of the system of acetonitrile for methyl alcohol afterwards
Survey.
That is, for methyl alcohol and the mixed system of acetonitrile, being mixed according to ratio as shown in the table
Using as solubilising reagent, and detected value (unit of the solubilising reagent of each ratio for TG is measured：
MM/l).
Table 1：The TG dissolution rates of different hemolyzing reagents
Similar, for methyl alcohol and the mixed system of acetonitrile, mixed according to ratio as shown in the table
Using as solubilising reagent, and the detected value of the solubilising reagent for TC of each ratio is measured.
Table 2：The TC dissolution rates of different hemolyzing reagents
Equally, for methyl alcohol and the mixed system of acetonitrile, mixed to make according to ratio as shown in the table
For solubilising reagent, and the detected value of the solubilising reagent for LDL-C of each ratio is measured.
Table 3：The LDL-C dissolution rates of different hemolyzing reagents
(note：Each value in above-mentioned table one to table three is the mean value of four parallel tests, and wherein * represents the significantly abnormal data that can be rejected.)
For HDL-C, above-mentioned all of different hemolyzing reagents are very low to its recovery rate, are presented as
The detected value for redissolving HDL-C in solution obtained using different hemolyzing reagents is significantly less than its corresponding whole blood
The detected value of serum, wherein the dissolved efficiency of HDL-C is most when by the use of 100% acetonitrile as hemolyzing reagent
Height, its detected value is the 40% of the detected value of corresponding WBS.Concentration for HDL-C, can be with
By other concentration levels of three, using Friedewald formula：TC=TG/5+HDL-C+LDL-
C is calculated and obtained.
It can be seen from data listed in table 1, table 2, table 3,100% methyl alcohol is compared as hemolyzing reagent,
The acetonitrile of 75% methyl alcohol+25% improves 45% to the recovery rate of TC, the recovery rate raising 9% to TG, right
The recovery rate of LDL-C improves more than 25%.
Additionally, according to data listed in table 1,2,3, it may be determined that the specific blood in different solubilising reagents
The correlation of the corresponding WBS detected value of the dry blood examination measured value of liquid composition.As a result show, 75% first
The dry blood examination measured values of TC and relative coefficient (the referred to as TC of TC Virus monitory values that the acetonitrile of alcohol+25% is extracted
As a result correlation) it is R2=0.6988 (Fig. 7 A), the correlation of TG results is R2=0.9423 (figure
7B), the correlation of LDL-C results is R2=0.4581 (Fig. 7 C).100% methyl alcohol is compared as molten
The TC that blood reagent is extracted is R with the correlation of serum TC result2=0.7101 (Fig. 8 A), TG results
Correlation be R2=0.7966 (Fig. 8 B), the correlation of LDL-C results is R2=0.1614 (figure
8C).As can be seen here the acetonitrile of 75% methyl alcohol+25% as hemolyzing reagent compared to pure methyl alcohol as hemolyzing reagent
When, the WBS detected value that the detected value of the specific blood constituent of its dry blood sample is corresponding has more preferably
Correlation.It should be noted that：Abscissa in Fig. 7 A-7C and Fig. 8 A-8C represents serum sample
Detected value (unit：MM/l), and ordinate then represents dry blood sample detected value (unit：MM/
Rise).
Embodiment 3：The background interference of different detection reagents
From data in the table 1-3 of embodiment 2, either for TC, TG or LDL-C, its
Dry blood sample when the mixed solution of methyl alcohol or methyl alcohol and acetonitrile is used as hemolyzing reagent, corresponding blood constituent
Detected value is all higher than its detected value in corresponding whole blood sample.That is, the preprocess method pair of blood sample is done
Detection reagent is answered to there is background interference.To detect whether that the interference is existed only in described in embodiment 1,2
Between the detection reagent of the Li Deman used in doing the preprocess method and embodiment 2 of blood sample, we purchase
Other commercially available TC, TG, LDL-C detection reagent (such as the last nine detections described in embodiment 1 are bought
Kit), and detected according to the content correspondence described in embodiment 2.Experiment find, with
100% methyl alcohol, the acetonitrile of 80% methyl alcohol+20%, the acetonitrile of 70% methyl alcohol+30% are being utilized not as hemolyzing reagent
When being detected with the detection reagent of company, there is interference in different hemolyzing reagent for detection reagent.
But, using TC, TG, LDL-C in the dry blood sample of Li Deman reagents detection, the detection of gained
Value than the detected value in the detected value that obtained using the detection reagent of the last nine and whole blood sample correlation more preferably,
And with less interference.
Embodiment 4：Hemolysis time and the selection of haemolysis temperature
Collection and preparation according to the sample described in embodiment 2, the pretreatment of dry blood sample and to specific blood
The method that liquid composition is detected, wherein for hemolysis time and haemolysis temperature are adjusted, to be determined to
At utmost dissolve optimal dissolution time and the temperature of specific blood constituent.Wherein adopt the second of 70% methyl alcohol+30%
Nitrile is hemolyzing reagent, the dry blood sample of each sample Jing different hemolysis time at a temperature of haemolysis, is finally determined
TC detected values be listed in the table below 4.
Table 4：TC dissolution rates under different hemolysis times or different solution temperatures
As shown in Table 4 data are visible, and lower 2 hours of room temperature is enough by the specific blood constituent in dry blood sample
TC dissolutions.In the range of identical dissolution time, under the conditions of room temperature with 37 DEG C, specific blood constituent
Dissolution efficiency has no significantly different, but carries out the correlation analysis table with Virus monitory value according to data in table 4
Bright, the detected value of its specific blood constituent of the dry blood sample of dissolving has more preferable with Virus monitory value under room temperature
Correlation (data do not include).
Also, different hemolysis times or the dissolution rate experiment of the TG under different solution temperatures and LDL-C
(data do not include) also indicates that, lower 2 hours of room temperature is enough by the TG and LDL-C in dry blood sample
Dissolution.Here is not repeated.
In this experiment, also carry out with 100% methyl alcohol or by hemolyzing reagent of the acetonitrile of 80% methyl alcohol+20%
Above-mentioned identical detection, as a result shows that preferred hemolysis time, haemolysis temperature are not in different hemolyzing reagents
There is the difference of conspicuousness, room temperature lower 2 hours equally can be effective in the corresponding experiment of latter two hemolyzing reagent
The specific blood constituent of ground dissolution, and condition of the room temperature compared with 37 DEG C can provide more preferable correlation (data not wrapped
Include).
Embodiment 5：The repeatability of specific blood constituent detection in pretreated dry blood sample
According to the collection and preparation of the sample described in embodiment 1, the pretreatment of dry blood sample and to specific blood
The method that composition is detected, wherein the total people of experimenter 5 (being represented with S1 '-S5 '), respectively takes whole blood sample
A and multiple dry blood cake pieces, wherein whole blood sample be stored in 4 DEG C at until use, and can be in being used for batch again
The dry blood cake piece of renaturation detection is sealed in the sample sack comprising drier and is stored in -80 DEG C or so until making
With, for batch between the dry blood cake piece of repeatable detection be sealed in the sample sack comprising drier and be stored in
20 DEG C or so until use.All dry blood samples use before detection 100% methyl alcohol (to examine as hemolyzing reagent
The toxicity of acetonitrile is considered, using 100% methyl alcohol as hemolyzing reagent in following experiment), in room temperature
2 hours of lower dissolving.
Detection is repeatable in batch
In single batch detection, 20 duplicate detections are carried out to same sample, by the repeated detection of each sample
As a result carry out averagely, and be calculated standard variance (SD) to be listed in the following table, wherein the coefficient of variation
(CV) represent that SD accounts for the percentage of mean value.
Table 5：Repeated detection TC value in batch
| Sample | N | Mean value | SD | CV |
| S1″ | 20 | 7.41 | 0.24 | 3.25% |
| S2′ | 20 | 6.24 | 0.22 | 3.57% |
| S3′ | 20 | 5.3 | 0.15 | 2.57% |
| S4′ | 20 | 5.59 | 0.2 | 3.62% |
| S5′ | 20 | 5.63 | 0.25 | 4.37% |
The CV of 5 sample TC detected values shown in table 5 is respectively：3.25%th, 3.57%, 2.75%,
3.62% and 4.37%.
Table 6：Repeated detection TG value in batch
| Sample | N | Mean value | SD | CV |
| S1″ | 20 | 1.15 | 0.05 | 4.25% |
| S2′ | 20 | 0.81 | 0.03 | 3.35% |
| S3′ | 20 | 0.73 | 0.03 | 3.49% |
| S4′ | 20 | 0.70 | 0.02 | 2.87% |
| S5′ | 20 | 1.28 | 0.05 | 3.84% |
The CV of 5 sample TG detected values shown in table 6 is respectively：4.25%th, 3.35%, 3.49%,
2.87% and 3.84%.
Table 7：Repeated detection LDL-C value in batch
| Sample | N | Mean value | SD | CV |
| S1″ | 20 | 4.47 | 0.17 | 3.82% |
| S2′ | 20 | 3.96 | 0.22 | 5.61% |
| S3′ | 20 | 3.30 | 0.09 | 2.8% |
| S4′ | 20 | 3.62 | 0.19 | 5.16% |
| S5′ | 20 | 3.07 | 0.11 | 3.61% |
The CV of 5 sample TG detected values shown in table 7 is respectively：3.82%th, 5.61%, 2.8%,
5.16% and 3.61%.
Detection is repeatable between batch
In 7 different detection batches, respectively duplicate detection is carried out to identical sample, by each sample
Repeated detection result is carried out averagely, and is calculated standard variance (SD) and CV is listed in the following table.
Table 8：Repeated detection TC value between batch
| Sample | N | Mean value | SD | CV |
| S1″ | 7 | 6.45 | 0.54 | 8.38% |
| S2′ | 7 | 5.51 | 0.46 | 8.34% |
| S3′ | 7 | 5.25 | 0.34 | 6.65% |
| S4′ | 7 | 5.76 | 0.40 | 6.91% |
| S5′ | 7 | 5.52 | 0.64 | 11.65% |
The CV of 5 sample TC detected values shown in table 8 is respectively：8.38%th, 8.34%, 6.65%,
6.91% and 11.65%.
Table 9：Repeated detection TG value between batch
| Sample | N | Mean value | SD | CV |
| S1″ | 7 | 1.20 | 0.15 | 12.89% |
| S2′ | 7 | 0.79 | 0.09 | 11.77% |
| S3′ | 7 | 0.79 | 0.08 | 9.57% |
| S4′ | 7 | 0.80 | 0.08 | 9.51% |
| S5′ | 7 | 1.39 | 0.11 | 7.65% |
The CV of 5 sample TG detected values shown in table 9 is respectively：12.89%th, 11.77%,
9.57%th, 9.51% and 7.65%.
Table 10：Repeated detection LDL-C value between batch
| Sample | N | Mean value | SD | CV |
| S1″ | 7 | 4.01 | 0.41 | 10.24% |
| S2′ | 7 | 3.64 | 0.37 | 10.15% |
| S3′ | 7 | 3.43 | 0.32 | 9.26% |
| S4′ | 7 | 3.86 | 0.26 | 6.81% |
| S5′ | 7 | 3.26 | 0.23 | 7.18% |
The CV of 5 sample TG detected values shown in table 10 is respectively：10.24%th, 10.15%,
9.26%th, 6.81% and 7.18%.
In above-mentioned batch and batch between duplicate detection test result indicate that, for the dry blood sample of Jing special pre-treatments
In specific blood constituent be particularly the detection of TC, TG, LDL-C there is extraordinary repeatability, can
See of the present invention practical for the detection method of specific blood constituent in dry blood sample.
Embodiment 6：The accuracy of specific blood constituent detection in pretreated dry blood sample
According to the collection and preparation of the sample described in embodiment 1, the pretreatment of dry blood sample and to specific blood into
The method for point being detected, wherein the total people of experimenter 33, respectively takes that whole blood sample is a and multiple dry blood cake pieces,
Dry blood cake piece is wherein directed to, before detection or punching or is sheared and is obtained dry blood sample.All dry blood samples are in detection
Before use 100% methyl alcohol as hemolyzing reagent, 2 hours are dissolved at room temperature.
The preparation of standard items：
After the mixing of the blood of multiple blood donors, upper plasma and lower floor deposition will be obtained after centrifugation
Thing.By TC, TG, LDL-C standard solution of above-mentioned deposit and commercially available concentration known (or Jing systems
There is TC, TG, LDL-C standard solution of different concentration knowns after row dilution) with 5.5:4.5 volume
Than mixing, the standard items for different specific blood constituents are obtained.
Specifically, 10ml whole blood samples are added in the centrifuge tubes of 15ml Nunc 339650, is centrifuged using LS
Machine Baiyang 400C are centrifuged, 3000rpm centrifugation 5min.After removal centrifugation on the about 4ml on upper strata
Clearly, after in centrifuge tube add 8ml PBS solutions (Sigma-Aldrich companies, article No.：P5368-10PAK),
Be gently mixed using pipettor, repeated centrifugation, 5 times the step of remove supernatant after obtain blood seabed sediment.
Blood fat Quality Control standard items Bio-Rads 641 of the 4ml from Bio-Rad companies is measured afterwards, and is added it to
It is gently mixed uniformly to obtain the first standard items in above-mentioned 15ml centrifuge tubes.Repeat the above steps, but use
Bio-Rad 642 replaces Bio-Rad 641, obtains the second standard items.And repeat the above steps, use other
The solution of the specific blood constituent with variable concentrations replaces Bio-Rad 642, Bio-Rad 641 to obtain n-th
Standard items.
25 the first standard items of μ l, the second standard items ... n-th standard items are taken, is instilled and is stored into
It is used to collect in the filter paper panel region of blood sample on the dry blood cake pieces of BM0401 so as to cover the area of whole filter papers,
Repeat the step 5 time point sample in 5 single blood sample collection regions for each standard items.Afterwards room temperature is done
Dry about 2 hours are until filter paper is parched.Dried dry blood cake piece is sealed in into the sample sack comprising drier
In, -80 DEG C are stored in until using.
Before detection above-mentioned dry blood cake piece is punched using punching instrument, or go out whole filters using scissor cut
Scraps of paper region is sampled to obtain one or more diameters between 1mm-10mm to the dry blood cake piece
Dry blood sample, the gross area sum of one or more dry blood samples of the standard items is equal to dry blood sample to be measured
Gross area sum.
The comparison of punching sampling mode and shearing sampling mode：
For the dry blood sample of standard items, using 100% methyl alcohol as hemolyzing reagent, at room temperature dissolving 2 is little
When.Redissolution solution is taken afterwards, detection reagent is added, afterwards according to the operating procedure and Amorsino of kit
The corresponding detection program of 460 automation biochemical instruments is detected that (what is wherein, taken answers to specific blood constituent
Solution and the increasedd volume of detection reagent must be consistent when detecting dry blood sample to be measured).
Meanwhile, 450 μ l the first standard items mentioned above, the second standard items ... n-th standard items are taken respectively,
Jing is centrifuged, and takes 150 μ l supernatants, adds detection reagent, afterwards according to the operating procedure of kit (the last nine) with
And the corresponding detection program of the automation biochemical instruments of Amorsino 460 is detected to specific blood constituent.
As a result show, the measured value of the specific blood constituent obtained according to WBS detection is with it according to theoretical
Calculated actual value (such as the concentration of each blood constituent is multiplied by 45% in Bio-Rad 641/642) is basic
Zero deflection.
It is special in the first standard items, the dry blood sample of the corresponding standard items of the second standard items ... n-th standard items
The detection of blood constituent is determined, using its result as the longitudinal axis, by the actual value of specific blood constituent in correspondence standard items
(serum true value) maps as transverse axis, and obtains calibration curve such as Fig. 9-10 institutes according to the trend between data point
Show.
Calibration curve wherein shown in Fig. 9 A-9C is that (by punching, instrument takes eight diameters by drilling method
The about region containing blood of 3mm) obtain when being sampled to the dry blood sample of standard items.Wherein as illustrated,
The calibration curve of TC is y=0.5834x+2.7557, wherein R2=0.9671；The calibration curve of TG is y=
0.8363x+0.1622, wherein R2=0.9729；The calibration curve of LDL-C is y=0.6171x+1.6813,
Wherein R2=0.9763.
Calibration curve shown in Figure 10 A-10C is that (three diameters of shearing are about 10mm by cutting method
Region containing blood) obtain when being sampled to the dry blood sample of standard items.Wherein as illustrated, the calibration of TC
Curve is y=0.6143x+3.8064, wherein R2=0.9973；The calibration curve of TG is y=0.9732x+
0.4463, wherein R2=0.9863；The calibration curve of LDL-C is y=0.5175x+2.8873, wherein R2
=0.9959.It should be noted that：Abscissa in Fig. 9 A-9C and Figure 10 A-10C represents serum sample
This detected value (unit：MM/l), and ordinate then represents dry blood sample detected value (unit：Mmoles
You/liter).
So, it can be seen from the correlation analysis of above-mentioned DBS samples and the detected value of serum sample, no matter
It is that the coefficient correlation of above-mentioned calibration curve is all higher than by shearing sampling mode or by the sampling mode that punches
96%, and the mean relative deviation of TC is respectively less than the mean relative deviation of 18%, TG and is respectively less than 29%；And
The mean relative deviation of LDL is respectively less than 23%.
Finally, can test according to more than, it is determined that calibration formula.Wherein direct comparison method is that industrial quarters is commonly used
Calibrating method, only directly compares sample and calibration curve, draws sample concentration.Calibration equation is typically simple
Linear equation, slope is immediately arrived at intercept by equation, is originated without going into slope and intercept.
Calibration formula
It is as follows for the calibration formula of " punching of DBS standard items punching+sample ":
TC:Y=x × A1-B1
Wherein y is result after the calibration of TC, the chemical result that x is measured for DBS samples, A1For calibration curve
Slope (in the range from 5.3-6.3), B1For calibration curve intercept (in the range from 3.8-4.8).
TG:Y=x × A2-B2
Wherein y is result after the calibration of TG, the chemical result that x is measured for DBS samples, A2For calibration curve
Slope (in the range from 4.1-5.1), B2For calibration curve intercept (in the range from 0.1-0.5).
HDL-C:Y=x × A3-B3
Wherein y is result after the calibration of HDL, the chemical result that x is measured for DBS samples, A3It is bent for calibration
Line slope (in the range from 4.4-5.3), B3For calibration curve intercept (in the range from 0.3-1.3).
LDL-C:Y=x × A4-B4
Wherein y is result after the calibration of LDL, the chemical result that x is measured for DBS samples, A4It is bent for calibration
Line slope (in the range from 5.2-6.5), B4For calibration curve intercept (in the range from 2.1-3.3).
And if when the sample volume that adopts of DBS standard items and sample is inconsistent, needing the base in above-mentioned equation
Volume calibration is carried out on plinth using volume correction factor.I.e. by the way that the x values are multiplied by into Vt/VsIt is (wherein described
The volume of included dry blood is V in dry blood sample to be measuredt, it is included in the dry blood sample of the standard items to do
The volume of blood is Vs) carrying out the volume correction.
In sum, it is special in above-mentioned method, the dry blood sample of detection for processing dry blood sample that the present invention is provided
Determine the detection method of blood constituent, it is possible to use high precision test equipment easily detects various composition in blood
The content of (such as four items of blood lipid tests), solving blood sampling difficult, sample transport inconvenience and holding time short etc. asks
Topic.
Also, the present invention proposes a kind of hemolyzing reagent that is quick, convenient, being easy to preparation, and by experiment
Demonstrate the specific blood constituent that hemolyzing reagent proposed by the present invention can be effectively dissolved in dry blood so that whole
Sample making course simplifies, so that extensive sample preparation, detection are possibly realized.
Further, the invention provides a kind of inspection for above-mentioned dry blood sample and corresponding specific blood constituent
The preparation method of the standard items of survey, improves the degree of accuracy of whole experiment.
Finally, the detection method also for specific blood constituent in above-mentioned dry blood sample of the invention provides one kind
The preparation method of Calibration equation such that it is able to more quickly and accurately obtain the actual value of specific blood constituent.
Above-mentioned schematic diagram is illustrated just to the purpose of example, is not limitation of the present invention.One
In the case of a little, some of which module or device can be added as needed on or reduced.
If although it should be noted that be referred to the equipment for drying or sub-device of testing equipment in above-detailed,
But it is this divide be merely exemplary and it is not enforceable.In fact, experiment of the invention
Example, the feature and function of above-described two or more devices can embody in one apparatus.Instead
It, the feature and function of an above-described device can be to come concrete by multiple devices with Further Division
Change.
The those skilled in the art of those the art can be by research specification, disclosure and accompanying drawing
And appending claims, understand and implement other changes of the embodiment to disclosing.In claim
In, word " including " is not excluded for other elements and step, and wording " one ", " one " are not excluded for again
Number.In the practical application of invention, a part possibility perform claim multiple technologies cited in requiring are special
The function of levying.Any reference in claim should not be construed as the restriction of the scope to claim.
Claims (26)
1. a kind of to doing the method that blood sample is pre-processed, including：
Obtain the dry blood sample comprising specific blood constituent；
Hemolyzing reagent is added in the dry blood sample to obtain the mixed liquor of the dry blood sample and hemolyzing reagent；
And
The mixed liquor is placed into certain dissolution time under specific solution temperature to obtain the dry blood sample
Redissolve solution,
The hemolyzing reagent includes the first organic solvent and the second organic solvent, wherein first organic solvent
Can be different from second organic solvent, or first organic solvent can be with described second
Organic solvent is identical, and when first organic solvent is identical with second organic solvent,
First organic solvent and second organic solvent are not methyl alcohol.
2. method according to claim 1, wherein first organic solvent and the second organic solvent are selected respectively
From the following group：Ethanol, isopropanol, ether, petroleum ether, n-hexane, acetonitrile or above-mentioned any combination.
3. method according to claim 1, wherein the hemolyzing reagent further includes PBS.
4. method according to claim 1, wherein the hemolyzing reagent includes methyl alcohol and acetonitrile.
5. method according to claim 4, wherein the content range of acetonitrile is 5%- in the hemolyzing reagent
30%.
6. method according to claim 4, wherein content of the methyl alcohol in hemolyzing reagent in the hemolyzing reagent
Scope is 70%-95%.
7. method according to claim 1, wherein the dissolution time is 15 minutes to 3 hours.
8. method according to claim 1, wherein the solution temperature is room temperature or 37 DEG C.
9. method according to claim 1, wherein the dry blood sample is obtained by the following method：
Blood sample is collected and is stored on dry blood cake piece, and makes the blood sample cover the whole dry blood
Patch；And
The dry blood cake piece is sampled to obtain dry blood sample.
10. method according to claim 8, wherein being sampled to obtain or many to the dry blood cake piece
Dry blood sample of the individual diameter less than or equal to 10mm.
11. methods according to claim 9, wherein the sampling is to the dry blood cake by using punching instrument
Piece punching is carried out.
12. methods according to claim 9, wherein the sampling is by the dry blood cake Pian Hanxue areas
Domain carries out what all shearings were carried out.
13. methods according to claim 11, wherein the dry blood cake piece is the dry blood scraps of paper.
14. methods according to claim 1, further include：Using mixed liquor described in mixed instrument waggle
Specific blood constituent in fully to dissolve the dry blood sample.
A kind of 15. methods of the standard items for preparing specific blood constituent, including：
Z random blood sample is obtained, and carries out mixing centrifugation with isolated upper plasma and seabed sediment,
Wherein Z is more than or equal to 3；
Obtain the standard liquid of the described specific blood constituent with known content；And
The deposit is mixed so that the standard items of the specific blood constituent are obtained with the standard liquid, wherein
Specific blood component concentration is R in the standard itemss。
16. methods according to claim 15, wherein by the deposit with the standard liquid with 5.5:4.5
Ratio mixing be obtained the specific blood component concentration as RsStandard items.
A kind of 17. methods for being detected to the specific blood constituent in dry blood sample to be measured, including：
The redissolution solution of the to be measured dry blood sample prepared according to method described in claim 1 is obtained, wherein described
The volume of included dry blood is V in dry blood sample to be measuredt；
Take VtdThe dry blood sample described to be measured of volume is redissolved solution and adds the detection of the specific blood constituent
Reagent is obtaining sample to be tested solution；
The sample to be tested solution is detected to obtain the detected value M of the specific blood constituent contentt；With
And
Using Calibration equation by the detected value M of the specific blood constituent contenttBe converted into the specific blood into
Divide actual value R of contentt。
18. methods according to claim 17, wherein the step of obtaining the Calibration equation is further included,
Including：
Obtain the R that N kinds contain different concentration knownss={ Rs1、Rs2、Rs3、Rs4…RsNSpecific blood constituent
The dry blood sample of standard items, included dry blood in the dry blood sample in wherein N >=5, and the standard items
Volume is Vs={ Vs1、Vs2、Vs3、Vs4…VsN}；
Obtain the redissolution solution of the dry blood sample of N kinds standard items prepared according to method described in claim 1；
Take VsdThe redissolution solution of the dry blood sample of the N kinds standard items of volume, and it is separately added into the specific blood
The detection reagent of liquid composition obtains N kind standard items sample solutions；
The N kinds standard items sample solution is carried out to detect the detected value for obtaining wherein specific blood constituent content
Ms={ Ms1、Ms2、Ms3、Ms4…MsN}；And
According to concentration known RsWith detected value MsBetween Relation acquisition normal equation.
19. methods according to claim 18, wherein the utilization Calibration equation is by the specific blood constituent
The detected value M of contenttIt is converted into actual value R of the specific blood constituent contenttProcess include：
By MtSubstitute in the Calibration equation as y values, calculate and obtain x values；And
By the way that x values are carried out into actual value R that volume correction obtains the specific blood constituent contentt。
20. methods according to claim 19, wherein working as Vtd=VsdWhen, by the way that the x values are multiplied by into Vt/Vs
To carry out the volume correction.
21. methods according to claim 19, wherein working as VtdIt is not equal to VsdWhen, by the way that the x values are taken advantage of
With (Vt/Vs)*(Vsd/Vtd) carrying out the volume correction.
22. methods according to claim 18, wherein further including to prepare containing concentration known RsIt is described
The step of standard items of specific blood constituent do blood sample, including：
Z random blood sample is obtained, and carries out mixing centrifugation with isolated upper plasma and seabed sediment,
Wherein Z is more than or equal to 3；
Obtain the standard liquid of the described specific blood constituent with known content；And
The deposit is mixed so that the standard items of the specific blood constituent are obtained with the standard liquid,
Specific blood component concentration is R in wherein described standard itemss。
23. methods according to claim 22, wherein by the deposit mix with the standard liquid with
5.5:4.5 ratio is to be obtained the specific blood component concentration as RsStandard items.
24. methods according to claim 22, wherein after the standard items of the specific blood constituent are obtained,
Further include：
The standard items point sample of the specific blood constituent and dry blood cake piece will be stored in, and covers the blood sample
The whole dry blood cake piece of lid；And
The dry blood cake piece is sampled a including V to obtainbThe standard items of the dry blood volume of volume are done
Blood sample.
25. methods according to claim 24, wherein the dry blood sample to be measured is to obtain by the following method
's：
Blood sample is collected and is stored on dry blood cake piece, and makes the blood sample uniform fold entirely described
Dry blood cake piece；And
The dry blood cake piece is sampled a including V to obtainsThe dry blood described to be measured of the dry blood volume of volume
Sample.
26. methods according to claim 1-25 any one, wherein the specific blood constituent is selected from the group：
T-CHOL, triglycerides, HDL-C, LDL-C or above-mentioned
Any combination.
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110160848A (en) * | 2019-06-06 | 2019-08-23 | 苏州博源医疗科技有限公司 | A kind of urine specimen pre-treating method |
| CN113884358A (en) * | 2021-10-09 | 2022-01-04 | 壹生检康(杭州)生命科技有限公司 | Preparation method of dried blood tablets |
Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2003052390A1 (en) * | 2001-12-17 | 2003-06-26 | Reclos J George | Simultaneous analysis of blood samples and estimation of blood (hemoglobin) content |
| CN1620512A (en) * | 2000-07-14 | 2005-05-25 | 爱科来株式会社 | Method of selectively determining glycated hemoglobin |
| EP1724587A1 (en) * | 2005-05-20 | 2006-11-22 | GermedIQ Forschungs- und Entwicklungsgesellschaft mbH | Method for determination of cardiovascular risk factors in dried blood |
| CN102159947A (en) * | 2008-08-21 | 2011-08-17 | 生物辐射实验室股份有限公司 | Calibration surface method for determination of analyte ratios |
| CN102323225A (en) * | 2011-06-09 | 2012-01-18 | 董理 | Method and reagent kit used for detecting low-density lipoprotein cholesterin |
| CN103760121A (en) * | 2014-02-18 | 2014-04-30 | 云南健牛生物科技有限公司 | Method for detecting nitrite in blood |
| CN204613163U (en) * | 2015-03-18 | 2015-09-02 | 杭州量康科技有限公司 | For detecting the system of dry blood sample |
-
2015
- 2015-10-14 CN CN201510661109.4A patent/CN106596209B/en active Active
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1620512A (en) * | 2000-07-14 | 2005-05-25 | 爱科来株式会社 | Method of selectively determining glycated hemoglobin |
| WO2003052390A1 (en) * | 2001-12-17 | 2003-06-26 | Reclos J George | Simultaneous analysis of blood samples and estimation of blood (hemoglobin) content |
| EP1724587A1 (en) * | 2005-05-20 | 2006-11-22 | GermedIQ Forschungs- und Entwicklungsgesellschaft mbH | Method for determination of cardiovascular risk factors in dried blood |
| CN102159947A (en) * | 2008-08-21 | 2011-08-17 | 生物辐射实验室股份有限公司 | Calibration surface method for determination of analyte ratios |
| CN102323225A (en) * | 2011-06-09 | 2012-01-18 | 董理 | Method and reagent kit used for detecting low-density lipoprotein cholesterin |
| CN103760121A (en) * | 2014-02-18 | 2014-04-30 | 云南健牛生物科技有限公司 | Method for detecting nitrite in blood |
| CN204613163U (en) * | 2015-03-18 | 2015-09-02 | 杭州量康科技有限公司 | For detecting the system of dry blood sample |
Non-Patent Citations (2)
| Title |
|---|
| NEAL E. CRAFT 等: "Retinol Analysis in Dried Blood Spots by HPLC", 《JOURNAL OF NUTRITION》 * |
| 谌敦丰: "溶血对干化学生化检验结果影响", 《医学影像与检验》 * |
Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110160848A (en) * | 2019-06-06 | 2019-08-23 | 苏州博源医疗科技有限公司 | A kind of urine specimen pre-treating method |
| CN113884358A (en) * | 2021-10-09 | 2022-01-04 | 壹生检康(杭州)生命科技有限公司 | Preparation method of dried blood tablets |
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