CN106148185B - Real-time electromagnetic field cell exposure system and its application - Google Patents
Real-time electromagnetic field cell exposure system and its application Download PDFInfo
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- CN106148185B CN106148185B CN201510177901.2A CN201510177901A CN106148185B CN 106148185 B CN106148185 B CN 106148185B CN 201510177901 A CN201510177901 A CN 201510177901A CN 106148185 B CN106148185 B CN 106148185B
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Abstract
Real-time electromagnetic field cell exposure system is made of C-shaped electromagnetic core, exciting current generator, fluorescence microscope, CCD camera, camera operation system, fluorescence image processing system, computer etc..C-shaped electromagnetic core is connect with exciting current generator and fluorescence microscope, fluorescence microscope is connect with CCD camera, CCD camera is connect with camera operation system, camera operation system is connect with exciting current generator, fluorescence image processing system and computer, and fluorescence image processing system is connect with computer.This real-time electromagnetic field cell exposure system is beneficial in that：While cell is by electromagnetic field exposure, the dynamic process that cell fluorescence changes over time is observed and recorded.To exclude possible endogenous or exogenous interference, accurate judgement cell electro-magnetic biological effect and its biophysical mechanism provide scientific research instrument.
Description
Technical field
The invention belongs to field of biotechnology, are related to a kind of cell fluorescence detection system, specifically, being related to one kind works as cell
When being exposed to electromagnetic field, with real-time detection and intracellular Fluorescence probe can be recorded to the system of electromagnetic field dynamic response.
Background technique
Electro-magnetic biological effect is reaction of the organism to electromagnetic field or electromagnetic wave, can be divided into fuel factor and non-thermal effect two
Kind.The fuel factor mechanism of action is clearer, is caused by Joule heat or radiation energy.Non-thermal effect is a kind of weak effect, acts on machine
It manages unclear, is the research hotspot of bioelectromagnetics.Electro-magnetic biological effect is a kind of connection reaction of cause and effect section, be initially electromagnetic field with
Physical reactions occur for biomolecule, and the biomolecule after reaction chemically reacts between each other, the biology after chemical reaction point
Son leads to cell effect, and cell effect leads to subsequent biologic-organ and whole reaction.Therefore, originally based on physical reactions
Effect is the starting point and key point of electro-magnetic biological effect.
The research of electro-magnetic biological effect has epidemiological survey, animal or human body, cell, molecule and biochemistry, biophysics etc.
Level.Currently, the research method of cell experiment compares between being mainly based upon the group observed after exposure, not in electromagnetic field exposure period
Between observe cell autoreactivity, i.e. non real-time nature.Thus, it may be difficult to confirm cytological effect caused by electromagnetic field, especially it is difficult
Other exogenous disturbing factors other than electromagnetic field are excluded, as cell sample removes moment by room temperature and incubator from incubator
Between the endogenous of stress reaction caused by the temperature difference and cell own metabolism interference etc..It is examined in real time in electromagnetic field exposure period
The biological effect for surveying cell, can reduce disturbing factor to the maximum extent, imitate to understanding unperturbed or low electromagnetic biological of disturbing under state
Characteristic is answered to be of great significance.
In cell experiment, active oxygen radical (ROS), intracellular Ca2+ (Ca2+), mitochondrial membrane potential (MMP) etc. be all
Important electro-magnetic biological effect observes object.ROS is the group or molecule (R.) with unpaired electronics, mainly there is superoxide anion
(), hydrogen peroxide (H2O2), hydroxy radical (.OH) etc., chemical property is active, and the service life is short, there is paramagnetism, is electromagnetism
The important symbol of biological originally effect.Intracellular Ca2+ (Ca2+) signal biography is played in the adjusting of cell function as second messenger
The key effect led is one of a variety of participation protein, phosphatide and activating molecules of enzyme of nucleic acid decomposition.Under normal circumstances, carefully
Calcium homeostasis intracellular is by plasma membrane Ca2+Translocase and the common manipulation and control of intracellular Ca2+ cell system.When cell damage, this manipulation
Process disorder can lead to Ca2+Interior stream increases, Ca intracellular2+Concentration uncontrollably continues to increase.Studies have shown that electromagnetic field exposure can
So that intracellular ion calcium concentration increases, and therefore, Ca2+It is the important symbol of signal transduction in electro-magnetic biological effect.Mitochondria is
Cellular energy production plant, the main place that ATP enzyme generates.Mitochondria generates ROS during breathing oxidation, in mitochondria
Film gathers proton and other ions and forms mitochondrial membrane potential (MMP).
Active oxygen radical (ROS), intracellular Ca2+ (Ca2+), mitochondrial membrane potential (MMP) can see with fluorescence probe
It examines.The fluorescence probe of ROS has DCFH-DA, Ca2+There are Fluo-3, MMP to have JC-1 etc..Fluorescent quenching refers to prolonging with the time
Long, fluorescent brightness gradually dimmed process, is a kind of fundamental characteristics of fluorescence.It is general thin due to the presence of fluorescent quenching characteristic
The detection technique of fluorescence of born of the same parents do not have in situ study, significantly the limit value in situ study of electromagnetic biological cytological effect.
A kind of all-in-one machine combined by electromagnetic field generator and fluorescence microimaging systems of the present invention is constituted
Real-time electromagnetic field cell exposure system, can be in electromagnetic field exposure period, the dynamic of observation in real time and record intracellular Fluorescence becomes
Change.
Summary of the invention
Real-time electromagnetic field cell exposure system be by C-shaped electromagnetic core 1, exciting current generator 2, fluorescence microscope 3,
The composition such as CCD camera 4, camera operation system 5, fluorescence image processing system 6, computer 7.C-shaped electromagnetic core 1 and excitation electricity
Flow-generator 2 and fluorescence microscope 3 connect, and fluorescence microscope 3 is connect with CCD camera 4, CCD camera 4 and camera operation
System 5 connects, and camera operation system 5 is connect with exciting current generator 2, fluorescence image processing system 6 and computer 7, fluorescence
Image processing system 6 is connect with computer 7.
C-shaped electromagnetic core 1 is made of silicon steel sheet heap 9 and magnet exciting coil 10.Silicon steel sheet heap 9 surrounds C-shape, and void is made
For cell slot 8, for placing cell ware.
Exciting current generator 2 is by signal generator 11, frequency regulator 12, current amplifier 13, exciting current tune
Section 14, exciting current switch 15, Hall sensor 16, Hall sensor detection circuit 17 form.Signal generator 11 and frequency
Adjuster 12 and current amplifier 13 connect, and current amplifier 13 is connect with exciting current adjustment 14 and exciting current switch 15.
Hall sensor 16 is connect with Hall sensor detection circuit 17.
Signal generator 11 generates the sinusoidal signal of 0-340Hz by frequency regulator 12, and is input to current amplifier
13.Current amplifier 13 generates the exciting current of 0-1A, 0-340Hz by exciting current adjustment 14, and is opened by exciting current
It closes 15 and is input to magnet exciting coil 10, make to generate magnetic induction intensity 0.01-25.37mT, the alternation of frequency 0-340Hz in cell slot 8
Magnetic field.Exciting current switch 15 controls water conservancy diversion and the blocking of exciting current, thus control in cell slot 8 generation of alternating magnetic field and
Stop.Hall sensor detection circuit 17 receives the magnetic field detection signal by Hall sensor 16, hands over for surveying in cell slot 8
The magnetic induction intensity of varying magnetic field realizes the quality control in magnetic field.
When exciting current generator 2 exports electric current, when cell slot 8 generates alternating magnetic field, cell ware is thin in cell slot 8
Born of the same parents are exposed in the magnetic field.Meanwhile fluorescence microscope 3 carries out microscopic imaging fluorescence to the cell, CCD camera 4 is aobvious to fluorescence
The image that micro mirror 3 is imaged is recorded in real time, is saved, and an image set arranged in temporal sequence is generated.Camera operation system
Unite 5 pairs of CCD cameras 4 unlatching, close, image capture, measurement, processing, record, preservation, label etc. are controlled.
Exciting current generator 2 issues an opening and closing letter while occurring or blocking electric current, to camera operation system 5
Number, and mark in image set, it is used to indicate the time that alternating magnetic field opens or closes.
Fluorescence image processing system 6 obtains image set from camera operation system 5, carries out figure to the image in image set
Picture sampling, cellular localization, cell recognition, fluorescence are calculated, as the processing such as line conversion, curvature correction and analysis, to realize time sequence
The image set of column is converted to the cell fluorescence curve of time series.
Fluorescence image processing system 6 is to calculate 21, as line by image sampling 18, cellular localization 19, cell recognition 20, fluorescence
Conversion 22, curvature correction 23 form.Image sampling 18 is to turn the analog quantity image of microscopy in such a way that A/D is converted
It is changed to digital picture.The color of digital picture is described with three primary colors (red, green, blue) brightness degree, each pixel primary colors (red, green,
It is blue) brightness degree be a byte (1bit), as 255 grades.Cellular localization 19 is first in the first frame image of image set
It is middle to mark single or multiple target cells with one or more boxes, then, in the second frame and later image, in same side
Frame position carries out edge extracting and Fluorescence Intensity Assays to the target cell of label.Cell recognition 20 is the choosing in cell image
Green component is selected as Threshold Analysis parameter, when the green component of a certain pixel is less than threshold value (< 50), then the pixel is labeled as
0, it is represented as background, otherwise, 1 is labeled as, is represented as cellular regions part.Threshold value 50 is that artificial setting can be required according to analysis
's.Take the noise jamming of image the connected region for automatically selecting maximum area labeled as 1, as target cell, other labels
It is 0, as background noise.Fluorescence measuring and calculating 21 be positioning label be cell compartment calculate the total fluorescent brightness of cell be area
Fluorescence intensity C (λ) the average value S of each pixel in domain.As line conversion 22 is surveyed to each of image set target cell fluorescence
After calculating average strength S, curve, the fluorescence curve figure of formation are connected into chronological order.The method of curvature correction 23 is when setting
Between be t, picture-taken frequency Ts, tpThe moment is cut for electromagnetic field.(t≤t before electromagnetic field exposurep), first to interested mesh
Mark cell collection TnTime (about 1-10 minutes), there is n=Tn/TsA data, measured value are F (t).Being established according to these data should
Target cell fluorescent quenching linear fit equation：F1(t)=at+b (0 < t≤tp).(the t > t after electromagnetic field incisionp), to sense
The target cell of interest acquires TmThere is m=T in timem/TsA data, and rectify a deviation to measured value F (t)：Δ F=F (t)-F1
(t) (t > tp).Cell fluorescence correction function is after being rectified a deviation：
The invention has the beneficial effects that：
This system can be observed while cell is by electromagnetic field exposure and what record cell fluorescence changed over time moves
State process.To exclude possible endogenous or exogenous interference, accurate judgement cell electro-magnetic biological effect, research cell moves in real time
Step response and its biophysical mechanism for causing biological effect to occur provide scientific research instrument.
Detailed description of the invention
Fig. 1 is real-time electromagnetic field cell exposure system figure.
Fig. 2 is 1 schematic diagram of C-shaped electromagnetic core.
Fig. 3 is 2 schematic diagram of exciting current generator.
Fig. 4 is 6 schematic diagram of fluorescence image processing system.
Fig. 5 is real-time electromagnetic field cell exposure ROS timing diagram intracellular.
Fig. 6 is real-time electromagnetic field cell exposure Ca intracellular2+Timing diagram.
Fig. 7 is fluorescent quenching calibration curve.
Specific embodiment
The present invention is described in detail with reference to the accompanying drawing.
Embodiment 1：
Referring to Fig. 1, real-time electromagnetic field cell exposure system is shown by C-shaped electromagnetic core 1, exciting current generator 2, fluorescence
The composition such as micro mirror 3, CCD camera 4, camera operation system 5, fluorescence image processing system 6, computer 7.C-shaped electromagnetic core 1 with
Exciting current generator 2 and fluorescence microscope 3 connect, and fluorescence microscope 3 is connect with CCD camera 4, CCD camera 4 and camera shooting
Head operating system 5 connects, and camera operation system 5 and exciting current generator 2, fluorescence image processing system 6 and computer 7 connect
It connects, fluorescence image processing system 6 is connect with computer 7.
Referring to fig. 2, C-shaped electromagnetic core 1 is made of silicon steel sheet heap 9 and magnet exciting coil 10.Silicon steel sheet heap 9 surrounds C-shape,
Void is as cell slot 8, for placing cell ware.
The electric current of 0-1A can occur for exciting current generator 2, and be input in the magnet exciting coil 10 of C-shaped electromagnetic core 1, with
Generate magnetic induction intensity 0.01-25.37mT, the alternating magnetic field of frequency 0-340Hz.Fluorescence microscope 3 is to the cell in cell ware
Fluorescence carries out micro-imaging, and the image that fluorescence microscope 3 is imaged in CCD camera 4 is recorded in real time, saved, and generates one
The image set arranged in temporal sequence, camera operation system 5 to the unlatching of CCD camera 4, close, Image Acquisition, measurement,
Processing, record, preservation, label etc. are controlled.
Exciting current generator 2 issues an opening and closing letter while occurring or blocking electric current, to camera operation system 5
Number, and mark in image set, it is used to indicate the time that alternating magnetic field opens or closes.
Fluorescence image processing system 6 obtains image set from camera operation system 5, carries out figure to the image in image set
Picture sampling, cellular localization, cell recognition, fluorescence are calculated, as the processing such as line conversion, curvature correction, to realize the figure of time series
Image set is converted to time series cell fluorescence brightness curve.
Embodiment 2：
Referring to Fig. 3, exciting current generator 2 is by signal generator 11, frequency regulator 12, current amplifier 13, encourages
Magnetic current regulation 14, exciting current switch 15, Hall sensor 16, Hall sensor detection circuit 17 form.Signal generator
11 connect with frequency regulator 12 and current amplifier 13, and current amplifier 13 and exciting current adjustment 14 and exciting current switch
15 connections.Hall sensor 16 is connect with Hall sensor detection circuit 17.
Signal generator 11 generates the sinusoidal signal of 0-340Hz by frequency regulator 12, and is input to current amplifier
13.Current amplifier 13 generates the exciting current of 0-1A, 0-340Hz by exciting current adjustment 14, and is input to exciting current
Switch 15.Exciting current switch 15 controls water conservancy diversion and the blocking of exciting current, to control the generation of alternating magnetic field in cell slot 8
And stopping.Hall sensor detection circuit 17 receives the magnetic field detection signal by Hall sensor 16, for surveying in cell slot 8
The magnetic induction intensity of alternating magnetic field realizes the quality control in magnetic field.
Embodiment 3：
Referring to fig. 4, fluorescence image processing system 6 is surveyed by image sampling 18, cellular localization 19, cell recognition 20, fluorescence
Calculate 21, as line conversion 22, curvature correction 23 form.The main task of fluorescence image processing system 6 is real-time from CCD camera 4
In record, the image set saved, the fluorescent brightness of interested cell image is converted into numeric data in temporal sequence, and
The fluorescent brightness curve in settling time domain.
The image of CCD camera 4 is acquired image data with 1 frame/second sample frequency by camera operation system 5, will be regarded
The image of Yezhong is sampled with 1340 × 1004 high-resolution, these data are transferred to camera behaviour by USB interface
Make in system 5, stored with the format of consecutive image, carries out subsequent image processing.Image sampling 18 is in such a way that A/D is converted
The analog quantity image of microscopy is converted into digital picture.Three primary colors (red, green, blue) brightness etc. of the color of digital picture
Grade description, the brightness degree of each pixel primary colors (red, green, blue) are a byte (1bit), as 255 grades.
Cellular localization 19 is for determining interested target cell, and processing method is selection imaging clearly, and fluorescence is bright
It is bright, and surrounding is used as analysis target without the cell of adhesion object, is generally positioned manually and is automatically positioned two kinds of image processing methods
Method：
(1) it is automatically positioned：Multiple cells using edge detection, in automatic identification image.This objective convenience of method, number
It is automated according to processing, can quickly handle a large amount of image data；
(2) it is positioned manually：First single or multiple mesh are marked with one or more boxes in the first frame image of image set
Cell is marked, then, in the second frame and later image, the target cell to label carries out edge extracting in same box position
And Fluorescence Intensity Assays.
In fact, many cells adhesion is very common phenomenon, automatic positioning often will appear deviation, and intervention is positioned manually
The judgement of people, it is ensured that the cell positioned is individually without adhesion, is practicable method.
The effect of cell recognition 20 is to identify cell compartment in fluorescent image, and the part of cellular regions is labeled as
1, remaining is labeled as 0.The fact that it is green that implementation method, which is according to cell fluorescence, and background is black, carries out two for cell image
Value processing.Processing method is to select green component as Threshold Analysis parameter, when the green of a certain pixel in cell image
Component is less than threshold value (< 50), then the pixel is labeled as 0, is represented as background, otherwise, is labeled as 1, is represented as cellular regions part.
Threshold value 50 can require manually to set according to analysis.
CCD camera 4 is a kind of charge coupled device based on semiconductor, and image quality is made an uproar by temperature change etc.
Acoustic jamming and it is unintelligible, showing some in cell fluorescent images is not that the image of cell is also displayed as green, causes binaryzation
Multiple images afterwards are connected to phenomenon.Processing method is to automatically select the connected region of maximum area labeled as 1, as target cell,
Other labels are, as background noise.
Fluorescence measuring and calculating 21 be positioning label be cell compartment calculate the total fluorescent brightness of cell.Calculation method be through
The fluorescent image that Image Acquisition obtains is the data indicated with R, G, B (RGB) three-component, according to International Commission on Illumination
The RGB system of digital picture is converted to CIE-XYZ system by specification (CIE).Firstly, RGB system is converted to CIE-XYZ system
Three-component values：
Then, chromaticity coordinate x, y, z are converted：
Based on y chromaticity coordinate component, search CIE different wave length and chromaticity coordinate x, y, z corresponding relationship contrast table are found
With y chromatic component color wavelength λ the most matched.CIE-XYZ system give the corresponding ideal tristimulus values of different wave length λ and
Space chromacity coordinate value.It takes chromaticity coordinate x, y to establish rectangular coordinate system, x, y value for representing different wave length is marked
Obtain a horseshoe curve.
For the pixel RGB values arbitrarily provided, chromaticity coordinate x, y of calculating is not necessarily located exactly on horseshoe curve.
If any a point P, in wavelength of the nearest point wavelength value on horseshoe line as the pixel, as P point on horseshoe line 480nm
Point is nearest, therefore the wavelength of P point is exactly 480nm.After obtaining wavelength X, CIE-RGB color space is searched for, is searched corresponding with wavelength
Three-component values R (λ), G (λ), B (λ), and calculate fluorescence intensity C (λ)：
C (λ)=R (λ)+4.590G (λ)+0.060B (λ) (3)
Corresponding wavelength is calculated from the RGB data of pixel, calculation amount is very big, especially for 30 minutes height of continuous acquisition
Image in different resolution, runing time are very big.Therefore, we are for exhaustible RGB, first establish λ (x, y)=λ (R, G,
B) inquiry table can be quickly obtained λ for a certain pixel, substantially increase arithmetic speed by data base querying.
Total fluorescent brightness of cell is fluorescence intensity C (λ) the average value S of each pixel in the region of cell recognition：
Referring to figs. 5 and 6, as line conversion 22 is to calculate average strength to each of image set target cell fluorescence
After S, curve, the fluorescence curve figure of formation are connected into chronological order.Fig. 5 and Fig. 6 is the exposure of electromagnetic field hippocampal neuron respectively
Phase ROS intracellular and Ca2+The real-time timing diagram of fluorescence.
Traditionally, bioluminescence technology is imaged for single, and fluorescent quenching does not influence image acquisition.But in situ study
In, fluorescent image is that multiple imaging forms image set, and fluorescent quenching will persistently reduce fluorescent brightness, form imaging deviation, influence
As line is converted.Therefore, as the fluorescence curve that line is converted must be corrected.The method of curvature correction 23 is：
If the time is t, picture-taken frequency Ts, tpThe moment is cut for electromagnetic field.(t≤t before electromagnetic field exposurep), first
T is acquired to interested target cellnTime (about 1-10 minutes), there is n=Tn/TsA data, measured value are F (t).According to this
A little data establish the target cell fluorescent quenching linear fit equation：
F1(t)=at+b (0 < t≤tp) (5)
Here：F1It (t) is that fit equation target cell fluorescent brightness calculated value, a and b join for least square fitting equation
Number：
Here：WithThe respectively average value of cell fluorescence measured value F and time value t.
(the t > t after electromagnetic field incisionp), T is acquired to interested target cellmThere is m=T in timem/TsA data, and
It rectifies a deviation to measured value F (t)：
Δ F=F (t)-F1(t) (t > tp) (8)
Cell fluorescence correction function is after being rectified a deviation：
Referring to Fig. 7, the method for curvature correction 23 can obtain preferable effect.
Embodiment 4：
Studying real-time electromagnetic field exposure hippocampal neuron ROS and Ca intracellular2+Application.
0.1mg/ml poly-D-lysine (Poly-L-lysine, Sigma) coated cell ware of 0.5ml is used in super-clean bench
30min is inhaled and is abandoned, dries overnight spare.With 75% alcohol disinfecting SD rat, life is terminated, cerebral hippocampus tissue is taken, strips meninx
And blood vessel, it is placed in ice bath dissection liquid.After hippocampal tissue is shredded, it is transferred to 0.25% trypsin solution (Trypsin of 2ml
(1X), Life technologies) in, 37 DEG C of digestion 20min.It inhales and abandons pancreatin, plant liquid termination with 2ml kind in 37 DEG C of warm bath and disappear
Change, then is washed hippocampal tissue 2 times with plantation liquid.2m1 kind is added in hippocampal tissue and plants liquid, is gently blown and beaten, is used with 1ml liquid-transfering gun
500 turns/min is centrifuged 30s, and cell suspension is made.After cell count, with plantation liquid diluting cells suspension, and press 1 × 106/ ml is close
Degree is seeded in cell ware 0.5ml.Liquid is changed using liquid full dose with Neurobasal-A after 4h, is used with Neurobasal-A within the 3rd day
Liquid half, which is measured, changes liquid, and adds 0.5 μ l, 10 μM of cytarabines (Ara-c, Sigma), then used liquid with Neurobasal-A every 3 days
Half amount changes liquid.Take the 7th day cell for subsequent experimental.
Before system uses this system, the Hall sensor 16 on exciting current generator 2 is inserted into the thin of C-shaped electromagnetic core 1
In born of the same parents' slot 8, exciting current is adjusted by frequency regulator 12 and exciting current adjustment 14, makes the magnetic induction intensity in cell slot 8
It is continuously adjusted in 0-340Hz, 0.01-25.37mT, to guarantee the qualification in magnetic field in cell slot 8.
System is in use, cell ware is placed in the cell slot 8 in C-shaped electromagnetic core 1.When exciting current generator 2 is defeated
Electric current out, when cell slot 8 generates alternating magnetic field, the cell of cell ware is exposed in the magnetic field in cell slot 8.Meanwhile fluorescence
Microscope 3 carries out microscopic imaging fluorescence to the cell, realizes the electromagnetic field exposure in C-shaped electromagnetic core 1 and fluorescence microscope 3
Fluorescence imaging is synchronous to be generated, and real-time purpose is reached.
After system use, the deduced image collection from camera operation system 5 of fluorescence image processing system 6, by turning as line
It changes, obtains the real-time electromagnetic field exposure curve of time series.
Embodiment 5：
Two exciting current generators 2 are had developed, Zhejiang Province Measure Science & Technology Research Institute is sent to detect, testing result such as 1 He of table
Table 2.Hall sensor voltage in cell slot：1-200mV；Magnetic induction intensity：0.01047-25.3679mT；Precision：1.623%；
Field frequency：0-340Hz.
1 1# machine magnetic signature of table
2 2# machine magnetic signature of table
Claims (4)
1. a kind of real-time electromagnetic field cell exposure system, the system is by C-shaped electromagnetic core (1), exciting current generator
(2), fluorescence microscope (3), CCD camera (4), camera operation system (5), fluorescence image processing system (6), computer (7)
Composition；C-shaped electromagnetic core (1) is connect with exciting current generator (2) and fluorescence microscope (3), fluorescence microscope (3) and CCD
Camera (4) connection, CCD camera (4) are connect with camera operation system (5), camera operation system (5) and exciting current
Generator (2), fluorescence image processing system (6) and computer (7) connection, fluorescence image processing system (6) are connect with computer (7)；
Wherein the C-shaped electromagnetic core (1) is made of silicon steel sheet heap (9) and magnet exciting coil (10), and silicon steel sheet heap (9) surrounds C-shape,
Its void is as cell slot (8), for placing cell ware；Exciting current generator (2) is by signal generator (11), frequency
Adjuster (12), current amplifier (13), exciting current adjustment (14), exciting current switch (15), Hall sensor (16), suddenly
You form sensor detection circuit (17)；Signal generator (11) is connect with frequency regulator (12) and current amplifier (13),
Current amplifier (13) is connect with exciting current adjustment (14) and exciting current switch (15)；Signal generator (11) passes through frequency
Adjuster (12) generates the sinusoidal signal of 0-340Hz, and is input to current amplifier (13)；Current amplifier (13) passes through excitation
Current regulation (14) generates the exciting current of 0-1A, 0-340Hz, and is input to magnet exciting coil by exciting current switch (15)
(10), make to generate magnetic induction intensity 0.01-25.37mT, the alternating magnetic field of frequency 0-340Hz in cell slot (8)；Exciting current is opened
Close water conservancy diversion and the blocking of (15) control exciting current；Hall sensor (16) is connect with Hall sensor detection circuit (17), suddenly
Your sensor detection circuit (17) receives the magnetic field detection signal by Hall sensor (16), hands over for surveying in cell slot (8)
The magnetic induction intensity of varying magnetic field.
2. real-time electromagnetic field cell exposure system according to claim 1, which is characterized in that the CCD camera (4)
Generate and record one image set arranged in temporal sequence, exciting current generator (2) occur or block electric current while,
A keying signal is issued to camera operation system (5), and is marked in image set, alternating magnetic field is used to indicate and opens or close
The time closed.
3. real-time electromagnetic field cell exposure system according to claim 1, which is characterized in that the fluorescence image processing
System (6) be by image sampling (18), cellular localization (19), cell recognition (20), fluorescence measuring and calculating (21), as line conversion (22),
Curvature correction (23) composition；The Red Green Blue brightness degree of each pixel of digital picture of image sampling (18) is one
Byte, as 255 grades；Cellular localization (19) is first single with one or more box labels in the first frame image of image set
A or multiple target cells, then, in the second frame and later image, same box position to the target cell of label into
Row edge extracting and Fluorescence Intensity Assays；Cell recognition (20) is to select green component as threshold parameter in cell image,
When the green component of a certain pixel is less than the threshold value that numerical value is 50, then the pixel is labeled as 0, represents background, otherwise, is labeled as 1,
Represent cellular regions part；Threshold value is manually set in the range of being not more than 50；The noise jamming of image is taken and is automatically selected most
The connected region of large area is labeled as 1, and as target cell, other labels are, as background noise；Fluorescence measuring and calculating (21) be
The cell compartment that the label of positioning is calculates fluorescence intensity C (λ) the average value S that the total fluorescent brightness of cell is each pixel in region；
As line conversion (22) is after calculating mean value of fluorescence intensity S to each of image set target cell, to connect in chronological order
At curve, the fluorescence curve figure of formation；Curvature correction (23) is t≤t before electromagnetic field exposurep, first thin to interested target
Born of the same parents' acquisition, is established according to these data in 0 < t≤tpIt is interior, target cell fluorescent quenching linear fit equation F1(t)=at+b,
WhereinWithRespectively cell fluorescence measured value F's and time value t is flat
Mean value；The t > t after electromagnetic field incisionp, rectify a deviation to interested target cell measured value F (t)：Δ F=F (t)-F1(t),
Cell fluorescence correction function is after being rectified a deviation：
4. real-time electromagnetic field cell exposure system according to claim 1 is studying real-time electro-magnetic biological effect and its mechanism
In application.
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