CN109142501A - Full-automatic polypeptide extracts flight time mass spectrum detector - Google Patents
Full-automatic polypeptide extracts flight time mass spectrum detector Download PDFInfo
- Publication number
- CN109142501A CN109142501A CN201810860788.1A CN201810860788A CN109142501A CN 109142501 A CN109142501 A CN 109142501A CN 201810860788 A CN201810860788 A CN 201810860788A CN 109142501 A CN109142501 A CN 109142501A
- Authority
- CN
- China
- Prior art keywords
- sample
- bar magnet
- protein
- crane
- crawler belt
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Abstract
The present invention provides a kind of full-automatic polypeptide that high-volume sample can extract simultaneously extraction flight time mass spectrum detector.The mass spectrograph includes: polypeptide extraction purification device, liquid relief liquid distributing device, automatic sample application device, mass spectrograph.The mode that the present invention is covered using electromagnetic wand and bar magnet, enables sample to be completely separated；And polypeptide chip is prepared by automatic sample application device；By mass spectrometric laser sampling, polypeptide mass spectrogram is obtained.Detector time-consuming provided by the invention is less, high-efficient, consistency is good, avoid sample between cross contamination, the existing mass spectrometric technical effect that can reach and be manually operated.
Description
Technical field
The present invention relates to body fluid albumen extraction and the automated systems of mass spectrum point sample, extract in body fluid in conjunction with paramagnetic particle method
Albumen realizes the automatic operation of mass spectrum chip point sample, to be used to prepare biochip, and is adopted by mass spectrometric laser
Sample obtains the mass spectrogram and relevant information of albumen, can be used to prepare biochip, is consequently belonging to Protein Detection field.
Background technique
With the implementation and propulsion of the Human Genome Project, life science has entered the genome times afterwards comprehensively.At this
A epoch, the main study subject of life science are functional genomics, including Structural genomics research and proteome research
Deng.Although the genome for having multiple species now is sequenced, usually there is the function of more than half gene in these genomes
It can be unknown.Strategy employed in functional genome at present, as genetic chip, expressed sequence analyze (Serial
Analysis of gene expression, SAGE) etc., it is all from the viewpoint of mRNA in cell, with the proviso that carefully
The level of mRNA reflects the level of protein expression in born of the same parents.But the fact from DNA mRNA protein not fully in this way, exist
The regulation of three levels, i.e. transcriptional level control (Transcriptional control), translation skill regulation
(Translational control), level modulation (Post-translational control) after translation.From mRNA angle
Consider, actually only includes transcriptional level control, protein expression level can not be represented comprehensively.Experiment also turns out, organizes
The correlation of middle mRNA abundance and protein abundance is simultaneously bad, and for low abundance proteins, correlation is worse.It is heavier
It wants, the posttranslational modification of protein complexity, the subcellular localization of protein or migration, protein-protein interaction
Deng then can not almost judging from mRNA level in-site.Undoubtedly, protein is the executor of physiological function, is the straight of biological phenomena
Agent is met, change mechanism of the life under physiology or pathological conditions will be illustrated directly to the research of protein structure and function.
The existence form and mechanics of protein itself, such as posttranslational modification, protein-protein interaction and protein conformation
Problem is still relied on and is directly solved to albumen Quality Research.Although the special natures such as the changeability and diversity of protein are led
Cause protein research technology much more complicated than nucleic acid and much more difficult, but exactly these characteristics are participated in and affected whole
A life process.
Traditional mode studied single protein has been unable to satisfy the requirement of genome times afterwards comprehensively.This be because
Are as follows: (1) generation of biological phenomena is often multifactor impact, necessarily involves multiple protein.(2) ginseng of multiple protein
Be woven into network, or it is parallel occur, or in cascade cause and effect.(3) when executing physiological function, the performance of protein is more
It is sample, dynamic and substantially stationary constant not as genome.Therefore to have to the complicated activity of life comprehensive and deep
Understanding, must study protein in entirety, dynamic, the level of network.Therefore in the mid-90 in last century, state
A new branch of science-proteomics (Proteomics) is produced on border, it be with the presence of intracellular all protein and
Its manner is research object.It may be said that the development of proteome research is not only life science when entering Post genome
One of the milestone and the core content of genome times afterwards comprehensively life science in generation.The rise of proteomics is to technology
There are new demand and challenge.The research of protein group is substantially large-scale to protein progress parallel on a cellular level
Divide the analysis of variance, often to handle thousands of kinds of protein simultaneously.Therefore, development is high-throughput, highly sensitive, high accuracy
Investigative technique platform be present so that quite a while internal protein group research in main task.Currently in international albumen
In matter group investigative technique, the holoprotein component that usually can be used in cell or tissue carries out Proteomic analysis.It can also be into
Row sample is classified in advance, i.e., all protein in cell or tissue is merotomized using various methods, carry out albumen respectively
Matter group research.
Mass spectrometry mainly by the analysis of the mass-to-charge ratio of the ion to sample realize to sample carry out it is qualitative and
A kind of quantitative method.Basic principle is the electrification gas ion for converting sample to movement, presses mass-to-charge ratio (m/z) in magnetic field
Size separation simultaneously records.Mass spectrography is the analysis method being most widely used, it can be provided forms including sample element, nothing
Machine, organic and bioanalysis structure, the quantitative analysis of complex mixture, the analysis of surface of solids structure and composition, sample Central Plains
The isotope ratio of son.Initial mass spectrograph is mainly used to measure the atomic weight of element or isotope, with ion-optical theory
Development, mass spectrograph are continuously improved, and application range is also constantly expanding, and has been widely used in nothing to later period the 1950s
The measurement of machine compound and organic compound.Now, the footprint of mass spectral analysis is throughout the technical field of every subjects, in solid
The fields such as physics, metallurgy, electronics, space flight, atomic energy, the earth and cosmochemistry, biochemistry and life science have wide
Application.Mass-spectrometric technique is filled with new vitality in the application of life science, more mass spectrographic develop, and forms unique
Tree species for bio-energy source.
In recent years, with electrospray-mass spectrometry (eleetrosprayionization, ESI) and substance assistant laser desorpted
Biological mass spectrometry skill based on attached mass-spectrometric technique (matrix assisted laserdesortion/onizatio, MALDI)
Art, they have highly sensitive and high quality detection range, are up to accurate analyzing molecules amount tens of thousands of big to the biology of hundreds of thousands
Molecule (protein molecular) is possibly realized, so that the quick analysis of a large amount of protein molecular be made to become a reality.
Electrospray-mass spectrometry and matrix assisted laser desorption ionization mass-spectrometric technique are two rails for being born in the latter stage eighties
Ionization technique.The appearance of this two technologies makes traditional mass-spectrometric technique for being mainly used for small-molecule substance research that revolutionary character have occurred
Change.They have highly sensitive and high quality detection range, so that in pmol (10-12) the even water of fmol (10-15)
Accurately analyzing molecules amount is up to tens of thousands of large biological molecules to hundreds of thousands and is possibly realized on flat, so that mass-spectrometric technique be made really to walk
The research field of life science is entered, and has rapidly been developed.
Electrospray-mass spectrometry (ESI) be capillary exit apply a high voltage, generated high electric field make from
The liquid of capillary outflow is atomized into tiny charged drop, and as solvent evaporates, the charge density of droplet surface is gradually increased,
Last drop disintegration is a large amount of ions with one or more charges, causes analyte in the form of single charge or multiple-charged ion
Into gas phase.The characteristics of electro-spray ionization is to generate highly charged ion rather than fragment ion, drops mass-charge ratio (m/z)
As low as the range that most analytical instrument of quality can detect, thus greatly expand the analyst coverage of molecular weight, ion it is true
Real molecular mass can also be calculated according to mass-to-charge ratio and electric line number.The advantage of electrospray ionization mass spectrum be exactly it can be convenient with it is a variety of
Isolation technics is used in combination, if one matter of liquid combination (LC-MS) is to combine liquid chromatogram with mass spectrum and reach detection macromolecular complex
The purpose of matter.
The basic principle of matrix assisted laser desorption ionization mass-spectrometric technique (MALDI-TOF) is that analyte is dispersed in matrix point
In son and crystal is formed, when with laser irradiation crystal, by the energy that substrate molecule is absorbed via radiation, leads to energy accumulation
And rapid heat production, to make host crystal distil, matrix and analyte is caused to expand and enter gas phase.Matter caused by MALDAI
Spectrogram is mostly single charge ion, thus the quality of the ion and peptide and protein in mass spectrogram has-mono- corresponding relationship.MALDI
The ion of generation is often detected with time-of-flight detector, and theoretically, as long as the length of tof tube is enough, TOF detector can
The mass number of detection molecules is the not no upper limit, therefore MALDI-TOF mass spectrum is well suited for the research to protein, polypeptide.
However, traditional body fluid holoprotein purification is to be added to sample in the pipe containing magnetic bead and chemical reagent,
It is prepared by artificial mixing and washing, elution.Time-consuming, low efficiency, flux is low, consistency is poor for this method presence,
It is possible that there are the shortcomings such as cross contamination, extensive, large batch of protein extraction and purity analysis work can not be effectively carried out
Make, is gradually difficult to meet quick, accurate, large batch of protein extraction and mass spectrographic requirement.
Summary of the invention
In view of the above-mentioned problems, the present invention provides a kind of high-volume sample while extracting, and it is time-consuming less, high-efficient, consistency
Albumen that is good, avoiding cross contamination automates extraction system.The principle of the invention lies in: the mode covered using electromagnetic wand+bar magnet,
Protein extraction is completed using automatic manipulator to act, and the nucleic acid of extraction is then passed through respectively by dispenser point by automatic guide rail
It is attached in sample panel and sample panel is transferred on chip point sample instrument by pipettor, genetic chip is prepared by point sample instrument,
By mass spectrometric laser sampling, the mass spectrogram and relevant information of albumen are obtained.
Therefore, for the research purpose for realizing quick, direct, continuous analysis of protein, paramagnetic particle method and electromagnetic wand are combined
Method is extracted the advantages of protein isolate, DNA chip, and first purpose of the invention is to provide a kind of for protein spectrum analysis sample
Extract the automation mass spectrograph with the continuous operation process of mass spectrum point sample chip, comprising:
(1) protein extraction purification devices, comprising: stepping crawler belt 10, crane -- crawler belt attachment device 101, three-dimensional stepping
Motor 102, the mobile slide bar 103 of crane, bar magnet crane 104, bar magnet 105, bar magnet cover crane 106, bar magnet set 107, sample
Product plate 108, sample panel fixed station 110, the mobile slide bar 111 of sample panel fixed station；
Wherein, three-dimensional stepper motor 102 is mounted in crane-crawler belt attachment device 101；Bar magnet crane 104 and magnetic
Stick 105 is mounted on the surface of bar magnet set crane 106 and bar magnet set 107, so that bar magnet 105 can be properly inserted bar magnet set
In 108；The mobile slide bar 103 of crane uses sleeve-like manner, and bar magnet crane 104 is mounted on outer tube, and bar magnet covers crane 106
It is mounted on the inner tube, so that bar magnet 105 and bar magnet set 107 can be on all around under the action of three-dimensional stepper motor 102
In lower movement and 108 sample well of sample panel being properly inserted on sample panel fixed station 110, to avoid bar magnet by sample pollution and just
In extracting bar magnet out at any time, to realize the absorption of magnetic bead moment and be detached from bar magnet set.
In one embodiment, the sample panel fixed station is additionally provided with the left detent gear of sample panel fixing clamp 109, sample panel
Plate 113.Wherein, 4 sample panel fixing clamps 109 are installed respectively on four angles of sample panel fixed station 110, for sample panel
108 are fixed on sample panel fixed station 110；Sample panel block board 113 is used to determine the position of sample panel fixed station 109；Sample
Plate fixed station moves left and right the underface that slide bar 111 is mounted on three-dimensional stepping crawler belt 10, and connect with sample panel fixed station 110；
Sample panel fixed station moves left and right the left side that motor 112 is mounted on sample panel fixed station detent baffle 113, for that will contain extraction
The sample panel 108 of good sample is moved to right side and is dispensed by liquid relief liquid separation equipment.
In another embodiment, bar magnet 105 uses electromagnet, just loses magnetism after power-off, it is easier to separate bar magnet set
107 and magnetic bead.In a particular embodiment, bar magnet 105 and bar magnet set 107 using 3 row, 8 column modes or 6 row, 12 column mode (
Away from being determined by 108 pitch of holes of sample panel), 24 samples can be at most handled simultaneously；Sample panel 108 is mounted on sample panel fixed station
On 110, and in the underface of bar magnet set 108, so that bar magnet set 108 is accurately inserted into sample panel 108.In preferred embodiment party
It, can also be in the separately installed motor 112 of Far Left of slide bar 111, for the sample panel containing the sample extracted in case
108 move to right the lower section of liquid relief liquid distributing device 2, so as to liquid relief packing.
(2) liquid relief liquid distributing device, including pipettor -- crawler belt attachment device 201, three-dimensional stepper motor 202, crane are sliding
Bar 203, pipettor 204, sample collection plate 206；
Wherein, i.e., three-dimensional stepper motor 202 is mounted on pipettor -- in crawler belt attachment device 201, to drive lifting slide bar
203 move up and down；Pipettor 204 is mounted on lifting slide bar 203, and sample panel 108 is walked by the mobile slide bar of sample panel in three-dimensional
Into under the drive of motor 112, it is moved to the underface of pipettor 204, so that liquid relief syringe needle 205 is properly inserted in sample panel 108；
Sample collection frame 206 is fixed on the underface of stepping crawler belt, shifts the sample to come to receive pipettor 204.
In one embodiment, the three-dimensional stepping crawler belt can be identical three-dimensional step with protein extraction device (1)
Into crawler belt 10.
In another embodiment, it is additionally provided with the sample collection plate fixing clamp 207 for fixing collecting board.It is specific at one
In embodiment, wherein the liquid-transfering needle head 205 uses eight platoon modes, while 8 samples of packing can be at most shifted, and move
Liquid product can adjust freely according to actual needs, and stepping crawler belt 10 is arranged right below syringe needle rinse bath 32, is located at
The right side of sample collection frame can be cleaned by ultrasound.In a particular embodiment, pipettor 204 arranges channel using 8 rows 12
Sample injector, liquid transfer gun head 205 use metal type liquid droping head.
(3) automatic sample application device, including three-dimensional stepping crawler belt 301, three-dimensional stepping crawler belt 10-- three-dimensional stepping crawler belt 301 connect
Connection device 302, three-dimensional stepper motor 303, crane slide bar 304, crane 305, spotting needle 306,
Wherein, three-dimensional stepping crawler belt 301 with it is three-dimensional stepping crawler belt 10 is vertical is fixedly mounted on device 302, crane is sliding
Bar 304 is mounted on three-dimensional stepping crawler belt 301；Crane 305 is mounted on crane slide bar 304；Spotting needle 306 is mounted on liter
It drops on frame, and protein chip 307 is mounted on the underface of three-dimensional stepping crawler belt 10.
In one embodiment, i.e. X-direction movement is driven by stepper motor 303 in the horizontal direction, and is carried out in stepping
Realization is moved on band 10, mobile controlled by stepper motor 303 of front-rear direction, that is, Y direction is realized, in vertical direction, that is, Z axis side
To movement, is driven and realized by stepper motor 303.
In another embodiment, protein chip 307 is located in protein chip fixing groove 308, and is lacked by protein chip
Angle detent 309 and detent slot 308 are accurately positioned；Spotting needle 306 can be led to using 4 row, 3 column form arrangement (3*4 arrangement)
Over cleaning slot 32 is cleaned.
Protein extraction device 1, liquid relief liquid distributing device 2 and point sample in any of the above-described embodiment, in the mass spectrograph
Device 3 is distributed in the same horizontal line, and shares a set of stepping crawler belt 10, thus sequence complete sample separation, sampling, point sample with
And the automatic operation of cleaning and the drying of spotting needle.
Also in any of the above-described embodiment, needle cleaning device shares a set of, i.e. rinse bath 32 with liquid relief liquid distributing device,
Ultrasonic cleaner 322, ultrapure water rinse bath 323, waste liquid tank 324, dry slot including vial 321, for accommodating alcohol
325。
Also in any of the above-described embodiment, sample panel 108 uses 96 hole deep-well plates, and sample collection plate (206) preferentially selects
With 384 microwell plates, bar magnet set 107,105 specification of bar magnet match with sample panel 108, and stepper motor 102 drives bar magnet crane
104 and bar magnet cover crane 106, realize the combination of bar magnet 105 and bar magnet set 107 with separate, and generate vibration, mix sample
It closes uniform.
In a specific embodiment, three-dimensional stepper motor (X, Y, Z-direction) drives bar magnet crane 104 and magnetic
Stick covers crane 106 in tri- directions movements of X, Y, Z, to realize the extraction process of sample.In a more particular embodiment,
Sample panel 108 is fixed on sample panel fixed frame 109, and one or so propeller is connected to below sample panel fixed frame 109
On 110, under the drive of stepper motor 112, the sample panel 108 containing the sample extracted is moved to right side liquid relief liquid separation and is filled
Set 2 lower section.
In addition in any of the above-described embodiment, drive of the pipettor 204 in stepper motor (two X, Z-direction motors)
Under, the sample extracted is dispensed into sample collection plate 206 from the transfer of sample panel 108, liquid transfer gun head 205 uses 8 volley of rifle fires,
And match with sample panel 108, sample collection plate 206.In specific embodiments, rinse bath 32 includes two vials
321 (concentration is respectively 100% and 50%), it is dry for accommodating ultrasonic cleaner 322, the ultrapure water rinse bath 322 of alcohol
Room 325, waste liquid cylinder 324.In a more particular embodiment, rinse bath 35 further includes alcohol pump, ultrapure water pump, positive displacement pump etc..
In any of the above-described embodiment, spotting needle 306 is arranged using 3 row, 4 column mode or 1 row, 12 column mode arranges, excellent
First select metal type liquid droping head.Stepper motor (X, Y, Z-direction 3) moves in the X, Y, Z direction for controlling spotting needle 306,
To be accurately positioned and realize deposition process.
Sample extraction device 1, liquid relief liquid distributing device 2 and spot sample device 3 are all controlled by a host computer, can be passed through
Several programs are arranged to edit in the management software installed on computer, and the extraction, packing and point sample of sample is cooperated to work.
Second purpose of the invention is to provide using above-mentioned automation mass spectrograph, completes automation and extracts albumen, liquid relief point
Match and the method for Mass Spectrometer Method, step include:
(1) protein extraction purifies: sample is added in sample plate hole, and bar magnet 105 is inserted into bar magnet set 107, however
Sample liquid is stirred in hole together, magnetic bead is added then with binding protein, after collecting magnetic bead, washes, washs three times, elutes through supersalt
Afterwards, the protein sample of purifying is obtained；
(2) albumen liquid relief is distributed: sample panel 108 is by the mobile slide bar of sample panel, in the drive of three-dimensional stepper motor 112
Under, it is moved to the underface of pipettor 204, so that liquid relief syringe needle 205, which is properly inserted in sample panel 108, draws protein solution, so
Protein solution is transferred in the sample collection frame 206 for being fixed on the underface of stepping crawler belt by pipettor 204 afterwards；
(3) mass spectrum chip point sample: the sample after spotting needle 306 is cleaned and dried, on pipette samples collecting frame 206
Drop then carries out point sample on protein chip 307, and carries out Mass Spectrometer Method.
In another embodiment, in the case where purifying need not be extracted, by what is extracted by other modes
Sample, such as nucleic acid, microprotein polypeptide, are added in sample collection plate 206, then complete chip point sample by spot sample device 3,
And then realize Mass Spectrometer Method.
Also in one embodiment, in order to uniformly dispense liquid, liquid to be packed is pre-applied in sample panel 108,
The parameters such as packing volume are controlled by external host computer, liquid subpackage work is completed by liquid relief liquid distributing device 2.
Detailed description of the invention:
Fig. 1: protein extraction purification devices, liquid relief liquid distributing device and automatic sample application device side view；
Fig. 2: protein extraction purification devices, liquid relief liquid distributing device and automatic sample application device perspective view；
Fig. 3: protein extraction purification devices, liquid relief liquid distributing device and automatic sample application device top view；
Fig. 4: protein extraction purification devices；
Fig. 5: bar magnet, bar magnet set and its crane and sample panel practice operation diagram.
Fig. 6: liquid relief liquid distributing device；
Fig. 7: automatic sample application device.
Fig. 8: the internal structure enlarged drawing of rinse bath.
Fig. 9: spotting needle installation diagram.
Figure 10: the present invention automates mass spectrograph testing result figure.
Figure 11: existing mass spectrograph detects results of comparison figure.
Technical effect
1, the protein extraction purification devices in instrument are the protein molecular quilts to dissociate from sample solution in a closed environment
Special is adsorbed onto magnetic-particle surface, and the impurity such as nucleic acid are not adsorbed and stay in the solution.React certain time and then
It under magnetic fields, separates magnetic-particle with liquid, recycles particle (magnetic bead -- protein mixture), then be with elution
Obtain pure protein.The device can be reduced infection and the pollution probability of protein, reduces the waste of consumptive material, improves efficiency,
Manual operation is reduced, purification degree is improved, accurately completes Protein Extraction, prevent the pollution that manual operation may cause, prevent people
Member is infected by sample.
2, the Protein Extraction device in instrument, can by the instruction of external computer host, under the control of pre-set programs,
Sample is mixed, magnetic bead transfer, cleaning, the operation such as elution, pure protein example solution and disposable place can be obtained
Reason great amount of samples greatly improves work efficiency.
3, the Protein Extraction device in instrument, every layer of 96 deep-well plates have lid covering during extracting protein
And work, completely in cloche, which prevent the cross contaminations of drug.Telecontrol equipment uses helical localization machine to formulate completely
Level is true, makes it possible the small quantitative exercise of instrument.
4, the liquid relief liquid distributing device in instrument, can be quasi- by the instruction of external computer host, under the control of pre-set programs
Really sample is shifted and dispensed, also liquid transfer gun head is cleaned and dried, can both reduce the labor intensity of operator, also
Avoid cross contamination and manual operation bring error and the extraneous contamination of sample.
5, the mass spectrum spot sample device in instrument can be realized high-precision control point liquid and accumulate, high cleanliness control point liquid quality,
Efficiently complete large batch of sample point sample demand；Meanwhile by completing evenly distributing for sample in the case where computer program controls, avoid
Volumetric errors caused by manual operations and possible extraneous contamination, while improving the speed and efficiency of point sample.
6, this instrument is by reasonable engineering design, solely by Protein Extraction purifying, sample distribution and point sample mass spectrum three
Vertical experimentation is dexterously integrated into a manipulation instrument, is realized without cross-infection, sterile operating process, the instrument after integration
Device significantly reduces the labor intensity of operator, reduces fault and infection during manual operation, enhances albumen core
The safety of piece preparation process, validity accelerate the suitable automated process detected with analysis, are the research of molecular biology
Provide a kind of tool of modernization.
7, in addition, instrument after integration, rationally, space utilization rate is high, and all operating process are by soft for the placement of each device
Part control, intelligentized design are automatically performed the extraction purification process of protein, save the time and efforts of experimenter, make
Protein Extraction and Mass Spectrometer Method work become light, easy.
Specific embodiment
The present invention is further described with case study on implementation with reference to the accompanying drawing.But it is as described below, it is only of the invention preferable
Embodiment is not intended to limit the present invention in any form, and is implemented according to the technical essence of the invention to above
Any simple modification, equivalent change and modification made by example, all of which are still within the scope of the technical scheme of the invention.
Embodiment one: the extraction and movement assigning process (referring to fig. 4-6) of protein example in body fluid
1, reagent adding and sample
In 1 piece of 96 formula deep-well plates, sample -- magnetic bead -- is sequentially added in conjunction with liquid, sample -- magnetic bead -- is in conjunction with liquid, sample
Product -- magnetic bead -- are in conjunction with liquid, cleaning solution, cleaning solution, cleaning solution, cleaning solution, cleaning solution, cleaning solution, eluent, eluent, elution
Liquid, and sealed 96 formula deep-well plates with sealed membrane, it is placed in appropriate environment.Wherein the magnetic bead, in conjunction with liquid, cleaning solution,
The reagent in known magnetic bead-protein extraction reagent kit can be used in eluent.
The sample panel of good reagent will have been added by correct orientation, be fixed on sample panel fixed station 110.
2, setting method and parameter
Using external computer, method is set, takes an action name, such as " -- liquid separation -- point sample is extracted ", sets
Bar magnet covers insertion depth, and sample -- magnetic bead -- is in conjunction with liquid incorporation time, washing, elution time and bar magnet position and movement routine
Deng all setting rear store method, this method selected in operation.
3, it automatically extracts and purifies
Good all parameters to be placed, start to automatically extract protein.Bar magnet set crane 106 starts to move down at this time,
And be inserted into sample, vibration is generated to mix sample, and then bar magnet crane 104 rises, and leaves liquid, makes mixed sample
Product -- magnetic bead stands, sufficiently combines.Then, bar magnet frame 104, bar magnet stock 106 move down, and bar magnet 105 is inserted into bar magnet set 107
In, and be inserted into sample simultaneously, collect magnetic bead -- protein particulate.Then under the drive of horizontal step motor 112, successively exist
It is washed in cleaning solution 1 and cleaning solution 2.In washing process, 107 separation of bar magnet 105 and bar magnet set, bar magnet set 107 is stayed in a liquid,
And pico- vibration, it is then allowed to stand, makes cleaning solution and magnetic bead -- protein is sufficiently combined, and is carried out second and is washed.To wash twice
Terminate, bar magnet 105 is combined with bar magnet set 107, is inserted into the second pipe cleaning solution, collects magnetic bead -- protein particulate, and be transferred to
In eluent, protein is eluted from magnetic bead, during elution, bar magnet 105 is first separated with bar magnet set 107, magnetic
Pearl -- protein shakes in eluent with 107 separation of bar magnet set, bar magnet set 107 slightly, divides magnetic bead and protein sufficiently
From bar magnet 105 is inserted into bar magnet set 107 later, is inserted into eluent together, and magnetic bead is collected, and extraction process is completed.
4, extracting solution dispenses
Under the control of external computer program, the pushing away in horizontal propeller 114 of sample panel 108 containing the sample extracted
Under dynamic, it is moved to the underface of right side pipettor 204, pipettor 204 first in cleaning device 32, successively will by alcohol trough 321
100% alcohol, 50% alcohol are injected into ultrasonic cleaner, to complete clear process twice, are then done in hothouse 325
Dry, then pipettor 204 is moved to the top of sample panel 108, accurately draws the sample and volume preset, moves again to sample
206 top of product collecting board, is accurately squeezed into the hole preset.Complete liquid relief liquid separation circulation.In this way, all extracting solutions are turned
Shifting is dispensed into sample collection plate 206.
Embodiment two: sample packing
1, preparation
Sample to be packed is added in sample collection plate 206, is mounted on sample panel fixed station 110, and in stepping electricity
Under the drive of machine 112, it is moved to 113 position of right side detent slot.Sample collection plate 206 is fixed on corresponding position.
2, method parameter is arranged
In external computer program, an action name is taken, such as " sample packing ", then where packing sample needed for setting
Hole location and the corresponding hole location in sample collection plate, dispense volume, the parameters such as liquid relief speed, after good all parameters to be placed, protect
This method is deposited, selects this method in operation.
3, sample is dispensed
Corresponding method is selected, starts to dispense sample.Under the control of outer computer, pipettor 204 is moved to cleaning
Right above device 32, successively pass through the cleaning of 100% alcohol, 50% alcohol and ultrapure water, then the drying in hothouse 325
(referring to Fig. 8) then moves to the surface of sample panel 108, moved again to after accurate pipette samples sample collection plate 206 just on
Side, sample is got in corresponding hole location.Primary packing circulation is completed, after the completion of to be packed, seals sample with dedicated plastic packaging film
Collecting board 206.
Embodiment three: chip sample point sample (referring to Fig. 7-9)
Spot sample device first carries out the cleaning of spotting needle 306 under the control of external computer program.Point sample instrument 3 or point sample dress
It sets 3 to be moved to right above cleaning device 32, successively by 100% alcohol, 50% alcohol and ultrapure water cleaning, then in drying
It is dry in room 325, it is moved to the surface of sample collection plate 206 later, it is accurate to draw extracting solution and volume, it then moves to
It, will be on extracting solution exact point to setting target position right above chip 307.Complete a spotting cycle.In this way, by point in need
On extracting solution point to corresponding target position.It spontaneously dries at room temperature, is then followed by matrix.
It is added to configured matrix solution in matrix substrate, unloads sample collection plate 206, matrix substrate is fixed on sample
On the position of product collecting board 206.The step of according to point sample, covers the matrix of corresponding volume on chip.Complete point matrix work
After work, chip 307 spontaneously dries at room temperature, is directly used in machine later and does mass spectrum acquisition.
Example IV: mass spectral analysis
By the printing operation of spot sample device, treated sample is titrated to mass spectrum chip according to scheduled volume
On, at this point, chip is placed into mass spectrometric injection port, point sample program is set, starts point sample.By spectrometer analysis, matter
The results are shown in Figure 10 for spectrum.
Using the tandem mass spectrum system (autoflex III TOF/TOF) of German Brooker company, it is analyzed by mass spectrometry,
Testing result is as shown in figure 11.This instrument is mainly manual point sample, and operation expends the time and is easy to pollute.
As shown in Figure 10 and comparative diagram 11, self-reacting device of the invention, mass spectra peak obtained is single, no miscellaneous peak, with
Results of comparison figure is compared, no significant difference, shows automation mass spectrograph of the invention, can be obtained or existing more than manual operation
Mass spectrometric technical effect.
Claims (7)
1. one kind completes the method that albumen, liquid relief distribution and Mass Spectrometer Method are extracted in automation by automation mass spectrograph, wherein
The mass spectrograph includes protein extraction purification devices, liquid relief liquid distributing device, automatic sample application device, and step includes:
(1) pass through protein extraction purification devices and complete protein extraction purifying: sample being added in sample plate hole, and by bar magnet
(105) it is inserted into bar magnet set (107), however stirs sample liquid in hole together, magnetic bead is added then with binding protein, collects magnetic bead
Afterwards, after washing through supersalt, wash three times, eluting, the protein sample of purifying is obtained；Wherein,
Protein extraction purification devices, comprising: stepping crawler belt (10), crane -- crawler belt attachment device (101), three-dimensional stepper motor
(102), crane is mobile slide bar (103), bar magnet crane (104), bar magnet (105), and bar magnet covers crane (106), bar magnet set
(107), sample panel (108), sample panel fixed station (110), sample panel fixed station is mobile slide bar (111), and sample panel fixed station is left
Move right motor (112)；
Wherein, three-dimensional stepper motor (102) is mounted on crane -- in crawler belt attachment device (101)；Bar magnet crane (104) and
Bar magnet (105) is mounted on the surface of bar magnet set crane (106) and bar magnet set (107), so that bar magnet (10) can be inserted accurately
Enter in bar magnet set (108)；Crane mobile slide bar (103) uses sleeve-like manner, and bar magnet crane (104) is mounted on outer tube,
Bar magnet set crane (106) is mounted on the inner tube, so that bar magnet (105) and bar magnet set (107) are in three-dimensional stepper motor (102)
Under the action of can be moved in X, Y, Z-direction and be properly inserted sample panel (108) sample well on sample panel fixed station (110)
In, to avoid bar magnet by sample pollution and convenient for extracting bar magnet out at any time, to realize the absorption of magnetic bead moment and be detached from bar magnet set；
Wherein, the sample panel fixed station is additionally provided with the left detent baffle (113) of sample panel fixing clamp (109), sample panel；Sample panel
Fixed station mobile slide bar (111) is mounted on the underface of three-dimensional stepping crawler belt (10), and connect with sample panel fixed station (110)；
Sample panel fixed station moves left and right the left side that motor (112) is mounted on the left detent baffle (113) of sample panel, for that will contain extraction
The sample panel (108) of good sample is moved to right side and is dispensed by liquid relief liquid distributing device；
(2) complete the distribution of albumen liquid relief by liquid relief liquid distributing device: sample panel (108) passes through the mobile slide bar of sample panel fixed station
(111), under the drive that sample panel fixed station moves left and right motor (112), it is moved to the underface of pipettor (204), and is moved
Liquid device (204) carries out mobile adjustment in X, Z-direction by three-dimensional stepper motor (202), so that liquid relief syringe needle (205) is properly inserted
Protein sample is drawn in sample panel (108), then pipettor (204) protein sample is transferred to be fixed on stepping crawler belt just under
In the sample collection frame (206) of side；Wherein,
Wherein, i.e., three-dimensional stepper motor (202) is mounted on pipettor (204) -- in crawler belt attachment device (201), to drive lifting
Frame slide bar (203) moves up and down；Pipettor (204) is mounted on crane slide bar (203), and sample panel (108) passes through sample panel
Mobile slide bar is moved to the underface of pipettor (204) under the drive that sample panel fixed station moves left and right motor (112), and
Pipettor (204) carries out mobile adjustment in X, Z-direction by three-dimensional stepper motor (202), so that liquid relief syringe needle (205) is accurately inserted
Enter in sample panel (108)；Sample collection plate (206) is fixed on the underface of stepping crawler belt, is shifted with receiving pipettor (204)
The sample come；
(3) mass spectrum chip point sample is completed by automatic sample application device: after spotting needle (306) is cleaned and dried, draws sample
Protein sample drop on product collecting frame (206) then carries out point sample on protein chip (307), and carries out Mass Spectrometer Method；Its
In,
Automatic sample application device includes three-dimensional stepping crawler belt (301), is carried out for three-dimensional stepping crawler belt (301) to be mounted on three-dimensional stepping
The attachment device (302) of band (10), three-dimensional stepper motor (303), crane slide bar (304), crane (305), spotting needle
(306),
Wherein, three-dimensional stepping crawler belt (301) with it is three-dimensional stepping crawler belt (10) are vertical is fixedly mounted on attachment device (302), rise
Drop frame slide bar (304) is mounted on three-dimensional stepping crawler belt (301)；Crane (305) is mounted on crane slide bar (304)；Point
Sample needle (306) is mounted on crane, and protein chip (307) is mounted on the underface of three-dimensional stepping crawler belt (10), passes through
Three-dimensional stepper motor (303) control spotting needle (306) is moved on X, Y, Z, protein chip (307) is accurately positioned and realizes a little
Sample process.
2. method described in claim 1, wherein bar magnet (105) uses electromagnet, just lose magnetism after power-off, it is easier to separate
Bar magnet set (107) and magnetic bead.
3. method of any of claims 1 or 2, wherein the liquid relief syringe needle (205) uses eight platoons in liquid relief liquid distributing device
Mode, while 8 samples of packing can be at most shifted, and pipetting volume can be adjusted freely according to actual needs, and stepping
Crawler belt (10) is arranged right below syringe needle rinse bath (32), is located at the right side of sample collection frame, can be cleaned by ultrasound.
4. method as claimed in claim 3, wherein protein chip (307) is located in protein chip fixing groove (308), and passes through egg
White chip unfilled corner detent (309) and protein chip fixing groove (308) are accurately positioned；Spotting needle (306) uses 4 row, 3 column shape
Formula arrangement, and can be cleaned by syringe needle rinse bath (32).
5. method as claimed in claim 3, wherein the protein extraction purification devices (1) in mass spectrograph, liquid relief liquid distributing device (2) and
Automatic sample application device (3) is distributed in the same horizontal line, and shares a set of stepping crawler belt (10), so that sequence completes sample point
From, sampling, point sample and spotting needle cleaning and drying automatic operation.
6. method described in claim 4 or 5, wherein needle cleaning device shares a set of with liquid relief liquid distributing device, i.e., syringe needle is clear
Washing trough (32), it is ultrasonic cleaner (322), ultrapure water rinse bath (323) including vial (321), for accommodating alcohol, useless
Liquid bath (324), dry slot (325).
7. method of claim 6, wherein controlling protein extraction purification devices (1), liquid relief by identical host computer
Liquid distributing device (2) and automatic sample application device (3), and according to management software be arranged program, thus automatically complete the extraction of sample,
Packing works with point sample.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201810860788.1A CN109142501B (en) | 2015-03-17 | 2015-03-17 | Full-automatic polypeptide extraction time-of-flight mass spectrometry detector |
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201510116693.5A CN104977349B (en) | 2015-03-17 | 2015-03-17 | Full-automatic polypeptide extracts flight time mass spectrum detector |
| CN201810860788.1A CN109142501B (en) | 2015-03-17 | 2015-03-17 | Full-automatic polypeptide extraction time-of-flight mass spectrometry detector |
Related Parent Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510116693.5A Division CN104977349B (en) | 2015-03-17 | 2015-03-17 | Full-automatic polypeptide extracts flight time mass spectrum detector |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN109142501A true CN109142501A (en) | 2019-01-04 |
| CN109142501B CN109142501B (en) | 2020-06-05 |
Family
ID=54274060
Family Applications (2)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510116693.5A Active CN104977349B (en) | 2015-03-17 | 2015-03-17 | Full-automatic polypeptide extracts flight time mass spectrum detector |
| CN201810860788.1A Active CN109142501B (en) | 2015-03-17 | 2015-03-17 | Full-automatic polypeptide extraction time-of-flight mass spectrometry detector |
Family Applications Before (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201510116693.5A Active CN104977349B (en) | 2015-03-17 | 2015-03-17 | Full-automatic polypeptide extracts flight time mass spectrum detector |
Country Status (1)
| Country | Link |
|---|---|
| CN (2) | CN104977349B (en) |
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110044677A (en) * | 2019-05-31 | 2019-07-23 | 四川省科源工程技术测试中心 | A kind of processing unit for before total content of organic carbon measurement in hydrocarbon source rock |
| CN110327659A (en) * | 2019-07-19 | 2019-10-15 | 天津迪沃特生物电子科技有限公司 | A kind of extraction and purification method and device based on magnetic fiber material |
| CN111141814A (en) * | 2020-01-04 | 2020-05-12 | 山东大学齐鲁医院(青岛) | Kidney endocrine detection device based on protein map |
| TWI765296B (en) * | 2020-07-15 | 2022-05-21 | 致茂電子股份有限公司 | Magnetic bar module with multiple stages and apparatus for extracting nucleic acid having the same |
| CN115290903A (en) * | 2022-09-30 | 2022-11-04 | 北京诺禾致源科技股份有限公司 | High-flux preparation method and device of omics products |
| CN116190195A (en) * | 2023-04-26 | 2023-05-30 | 四川质谱生物科技有限公司 | Mass spectrometer detection reagent business turn over device and mass spectrometer |
| WO2023123571A1 (en) * | 2021-12-31 | 2023-07-06 | 博尔诚（北京）科技有限公司 | Fully-automatic nucleic acid methylation treatment device and treatment mechanism |
Families Citing this family (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN108931415B (en) * | 2017-05-23 | 2021-02-19 | 北京诚智光辉科技有限公司 | Cell film-making dyeing all-in-one with wiper mechanism |
| WO2020142798A1 (en) * | 2019-01-11 | 2020-07-16 | Meon Medical Solutions Gmbh & Co Kg | Pipetting device and method for the transfer of fluids |
| CN110093343A (en) * | 2019-05-08 | 2019-08-06 | 中国人民解放军军事科学院军事医学研究院 | A kind of integrated portable nucleic acid-extracting apparatus and method for extracting nucleic acid |
| CN111235639B (en) * | 2020-03-06 | 2021-04-20 | 通用生物系统（安徽）有限公司 | Phage antibody elution device |
Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2010021509A2 (en) * | 2008-08-21 | 2010-02-25 | Korea Research Institute Of Standards And Science | Quantification method of biochemical substances using ion scattering spectroscopy and specific-binding efficiency quantification method of biochemical substances using ion scattering spectroscopy |
| CN101776642A (en) * | 2010-02-04 | 2010-07-14 | 复旦大学附属中山医院 | Mass spectrometric detection method for ca-dependent secretory protein 1 in human serum |
| CN102492603A (en) * | 2011-11-22 | 2012-06-13 | 马庆伟 | Automated instrument for nucleic acid extraction and mass spectrum sample application |
| CN103197081A (en) * | 2013-04-17 | 2013-07-10 | 无锡优创生物科技有限公司 | Protein chip and preparation and detection method thereof |
| CN104411833A (en) * | 2012-06-26 | 2015-03-11 | 里尔大学地区医疗中心 | Method for the in vitro diagnosis of an invasive fungal infection by maldi-tof mass spectrometry |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1313622C (en) * | 2002-08-02 | 2007-05-02 | 赵翀 | High-flux cell biological chip testing technology and reagent case |
| JP4606270B2 (en) * | 2005-07-29 | 2011-01-05 | 株式会社神戸製鋼所 | Time-of-flight measurement device for sample ions, time-of-flight mass spectrometer, time-of-flight mass spectrometry method |
-
2015
- 2015-03-17 CN CN201510116693.5A patent/CN104977349B/en active Active
- 2015-03-17 CN CN201810860788.1A patent/CN109142501B/en active Active
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2010021509A2 (en) * | 2008-08-21 | 2010-02-25 | Korea Research Institute Of Standards And Science | Quantification method of biochemical substances using ion scattering spectroscopy and specific-binding efficiency quantification method of biochemical substances using ion scattering spectroscopy |
| CN101776642A (en) * | 2010-02-04 | 2010-07-14 | 复旦大学附属中山医院 | Mass spectrometric detection method for ca-dependent secretory protein 1 in human serum |
| CN102492603A (en) * | 2011-11-22 | 2012-06-13 | 马庆伟 | Automated instrument for nucleic acid extraction and mass spectrum sample application |
| CN104411833A (en) * | 2012-06-26 | 2015-03-11 | 里尔大学地区医疗中心 | Method for the in vitro diagnosis of an invasive fungal infection by maldi-tof mass spectrometry |
| CN103197081A (en) * | 2013-04-17 | 2013-07-10 | 无锡优创生物科技有限公司 | Protein chip and preparation and detection method thereof |
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110044677A (en) * | 2019-05-31 | 2019-07-23 | 四川省科源工程技术测试中心 | A kind of processing unit for before total content of organic carbon measurement in hydrocarbon source rock |
| CN110327659A (en) * | 2019-07-19 | 2019-10-15 | 天津迪沃特生物电子科技有限公司 | A kind of extraction and purification method and device based on magnetic fiber material |
| CN111141814A (en) * | 2020-01-04 | 2020-05-12 | 山东大学齐鲁医院(青岛) | Kidney endocrine detection device based on protein map |
| TWI765296B (en) * | 2020-07-15 | 2022-05-21 | 致茂電子股份有限公司 | Magnetic bar module with multiple stages and apparatus for extracting nucleic acid having the same |
| WO2023123571A1 (en) * | 2021-12-31 | 2023-07-06 | 博尔诚（北京）科技有限公司 | Fully-automatic nucleic acid methylation treatment device and treatment mechanism |
| CN115290903A (en) * | 2022-09-30 | 2022-11-04 | 北京诺禾致源科技股份有限公司 | High-flux preparation method and device of omics products |
| CN115290903B (en) * | 2022-09-30 | 2023-01-20 | 北京诺禾致源科技股份有限公司 | High-flux preparation method and device of omics products |
| CN116190195A (en) * | 2023-04-26 | 2023-05-30 | 四川质谱生物科技有限公司 | Mass spectrometer detection reagent business turn over device and mass spectrometer |
| CN116190195B (en) * | 2023-04-26 | 2023-06-27 | 四川质谱生物科技有限公司 | Mass spectrometer detection reagent business turn over device and mass spectrometer |
Also Published As
| Publication number | Publication date |
|---|---|
| CN104977349A (en) | 2015-10-14 |
| CN104977349B (en) | 2019-04-23 |
| CN109142501B (en) | 2020-06-05 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN104977349B (en) | Full-automatic polypeptide extracts flight time mass spectrum detector | |
| EP2633327B1 (en) | System layout for an automated system for sample preparation and analysis | |
| Elpa et al. | Automation of mass spectrometric detection of analytes and related workflows: A review | |
| CN102492603B (en) | Automated instrument for nucleic acid extraction and mass spectrum sample application | |
| US9207152B2 (en) | Pretreatment apparatus and mass analyzing apparatus equipped with the same | |
| US20020068366A1 (en) | Proteomic analysis by parallel mass spectrometry | |
| CN108459133B (en) | Automatic intelligent detection instrument for thin-layer chromatography analysis and real-time online detection method thereof | |
| CN1498129A (en) | Robotic autosampler for automated electrospray from microfluidic chip | |
| JP2003514226A (en) | Multi-dimensional electrophoresis device | |
| CN1675739A (en) | System and method for the preparation of arrays of biological or other molecules | |
| AU2015261711B2 (en) | Automated system for sample preparation and analysis | |
| CN106770814B (en) | A kind of protein chromatography separation platform and its application | |
| CN108878253A (en) | Mass spectrometric data acquisition method | |
| US20100273170A1 (en) | Method and device for preparing biological samples for analysis | |
| JP2013096733A (en) | Analytical device and analytical method | |
| US11879880B2 (en) | Integrated machines and methods for performing fully-automated biological evaluation and chemical analysis | |
| CN108445135A (en) | A kind of detection method for a variety of residue of veterinary drug rapid screenings | |
| Leduc et al. | Massively parallel sample preparation for multiplexed single-cell proteomics using nPOP | |
| JP7051871B2 (en) | Methods for Identifying Reagents During Processes in Analytical Systems | |
| Chen et al. | Combination of flow injection with electrophoresis using capillaries and chips | |
| CN212410248U (en) | Full-automatic nuclide solid phase extraction device | |
| Truong et al. | What’s new in single-cell proteomics | |
| WO2020129118A1 (en) | Mass spectrometer | |
| Wallman et al. | Robotic implementation of a microchip-based protein clean-up and enrichment system for MALDI-TOF MS readout | |
| Rossi | Parallel automation for chemical analysis |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| PE01 | Entry into force of the registration of the contract for pledge of patent right | ||
| PE01 | Entry into force of the registration of the contract for pledge of patent right |
Denomination of invention: Automatic time of flight mass spectrometer for peptide extraction Effective date of registration: 20210129 Granted publication date: 20200605 Pledgee: Beijing technology intellectual property financing Company limited by guarantee Pledgor: BEIJING CLIN BOCHUANG BIOTECHNOLOGY Co.,Ltd. Registration number: Y2021990000133 |
|
| PC01 | Cancellation of the registration of the contract for pledge of patent right | ||
| PC01 | Cancellation of the registration of the contract for pledge of patent right |
Date of cancellation: 20220314 Granted publication date: 20200605 Pledgee: Beijing technology intellectual property financing Company limited by guarantee Pledgor: BEIJING CLIN BOCHUANG BIOTECHNOLOGY Co.,Ltd. Registration number: Y2021990000133 |