CN104502254A - Automatic cell cloning survival rate detection device and detection method thereof - Google Patents
Automatic cell cloning survival rate detection device and detection method thereof Download PDFInfo
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Abstract
The invention provides an automatic cell cloning survival rate detection device which comprises a sample image acquisition instrument and an information processor, wherein the sample image acquisition instrument consists of five parts, namely a rectangular box, a CCD (Charge Coupled Device) camera, a three primary color light source, a fluorescent lamp light source and a push-pull tray; two grooves used for respectively accommodating a to-be-detected cell cloning sample and a control cell cloning sample are formed in the push-pull tray; the information processor is connected with the sample image acquisition instrument by virtue of a USB data line and comprises an information processing singlechip, an LCD screen, an SD interface and multiple function buttons. The invention also provides a detection method of the automatic cell cloning survival rate detection device. An automatic detection mode is adopted, the operation is simple and convenient, the detection error caused by a human mode is avoided, the detection precision and efficiency are greatly improved, the manual cost is saved, and the detection data is conveniently reviewed and traced by virtue of data archiving and saving functions.
Description
Technical field
The present invention relates to environment pollution detection technical field, specifically a kind of cell clone survival rate automatic detection device and detection method thereof.
Background technology
Along with the fast development of human social economy and the intensification day by day of industrialization degree, the physical environment that the mankind depend on for existence is but subject to more and more serious pollution, comprises atmospheric pollution, water pollutions, soil pollution and nuclear radiation etc.It is said, in worldwide, about have the sewage of billion cubic meter more than 4200 to enter rivers,lakes and seas every year, pollute the fresh water of 55000 billion cubic meters, account for more than 14% of global run-off.In China, the situation of the environmental pollutions such as current air, water source is very severe.Pollution entering the water comprises heavy metal ion in chemical products and poisonous organism, and these materials can cause people's deformity, cancer stricken, organ lesion, and serious threat is to the health of the mankind.How the toxicity of efficient detection pollutant is with effective monitoring with evaluate various environmental pollution and become one of important topic improving China's socio-economic development quality.
Environment pollution detection can physically based deformation, chemistry or Biological Principles, as Chinese invention patent CN101882350B discloses a kind of water pollution organism early warning system based on principle of microbial fuel cells and method, propose the variable quantity that detection water body enters the electric signal caused by microbiological fuel cell (MFC), carry out after process as calculated uploading to store also showing on a display screen in real time; Chinese invention patent application CN101692034A discloses a kind of portable on-line device for detecting water pollutants, diversion mode is adopted water sample to be measured to be directed to the district with a tight waist of Focused Gaussian Beam, measured mean grain size and the particle number concentrations of micron particles pollutant in water by transmission fluctuation method, use DSP to carry out Signal sampling and processing.
Compared with the water pollutions detection method of traditional physically based deformation or the principles of chemistry, detection method based on Biological Principles can reflect the potential cytotoxicity of environmental pollution more truly, there is the advantage of Reality simulation environmental pollution by water to biosome deleterious effect, therefore, [Ohe, T. is more and more subject to people's attention in recent years, Watanabe, T., and Wakabayashi, K. (2004) Mutagens in surface waters:a review.
mutat Res 567, 109-49].MTT colorometric assay and Cell clonality experiment are two kinds of important vitro cytotoxicity experiments.MTT colorometric assay exogenous MTT can be made to be reduced to water-insoluble bluish violet crystallization first a ceremonial jade-ladle, used in libation and the principle be deposited in cell based on the succinate dehydrogenase in living cells mitochondria, is mainly used in detection cytotoxicity experiment.And Cell clonality experiment is the characteristic based on living cells with normal proliferative, is mainly used in detection cell-proliferation activity, reflects the viability of cell better, therefore have more biological significance.
The water pollutions toxicity assessment system based on people's mouse hybridoma clonality that we have set up, water pollutions toxic degree is indicated by detecting cell clone survival rate, its detection perform by the limitation of toxic compounds kind in water environment, can not reflect the deleterious effect of poisonous potpourri on cell proliferation ability in aqueous medium better.Therefore, high precision, full-automatic cell clonal survival detect the active demand having become environmental pollution monitoring and the improvement comprising water pollutions.
Current, the mensuration of cell clone survival rate mainly adopts artificial counting mode: first carry out artificial counting to the cell clone in testing sample and control sample, then, by the Survival clone number of contrast testing sample and control sample, cell clone survival rate is carried out quantitatively.Usually, be intensively distributed with the individual cell clone (group) do not waited of hundreds of in a sample (Tissue Culture Dish), adopting artificial mode to carry out counting to these cell clones wastes time and energy.Meanwhile, due to the interference by subjective factors such as fatigue, individual character moods, manual type is adopted also to be difficult to ensure accuracy of detection.Prior, owing to can only carry out guestimate to clone's yardstick, manual type is difficult to distinguish the clone (group) meeting specific cells Active pharmaceutical, and then causes insecure cell clone survival rate to detect.
Summary of the invention
The object of the present invention is to provide a kind of cell clone survival rate automatic detection device and detection method thereof, digital image information scanning is carried out to cell clone sample to be measured and compared with control cells clone sample simultaneously, utilize image pattern recognition process and analysis of cells clone sample image, by the auto Segmentation of cell clone with identify the Aulomatizeted Detect realizing cell clone survival rate.
Technical scheme of the present invention is:
A kind of cell clone survival rate automatic detection device, described cell clone survival rate automatic detection device comprises sample image Acquisition Instrument and message handler, the push-and-pull pallet that described sample image Acquisition Instrument comprises rectangular box, is arranged on the ccd video camera of rectangular box inside top central authorities, is arranged on the three primary light source of rectangular box inside top surrounding, is arranged on the fluorescent light source of rectangular box bottom inside central authorities and is arranged in rectangular box; The sidepiece of described rectangular box adopts light barrier, and the central part of described push-and-pull pallet offers two grooves, is respectively used to place cell clone sample to be measured and compared with control cells clone sample;
Described message handler is connected with sample image Acquisition Instrument by USB data line, described message handler comprises information processing single-chip microcomputer, LCD display, SD interface and some function buttons, the output terminal of described function button is connected with the input end of information processing single-chip microcomputer, the output terminal of described information processing single-chip microcomputer respectively with three primary light source, fluorescent light source is connected with the input end of LCD display, described information processing single-chip microcomputer is connected with ccd video camera interactive mode, the output terminal of described information processing single-chip microcomputer is connected with the input end of SD card memory by SD interface.
Described cell clone survival rate automatic detection device, the both sides inwall of described rectangular box is respectively equipped with the grooved tracks for placing push-and-pull pallet.
Described cell clone survival rate automatic detection device, the cross section of described push-and-pull pallet is L-type, and its long leg is horizontally set in rectangular box, and its short leg is vertically arranged on rectangular box front end, and the central part of described short leg is provided with groove type handle.
Described cell clone survival rate automatic detection device, described three primary light source comprises the electronic color-changing control circuit board triphosphor tube light identical with four specifications, the input end of described electronic color-changing control circuit board is connected with the output terminal of information processing single-chip microcomputer, and the described output terminal of electronic color-changing control circuit board is connected with the input end of triphosphor tube light; Described fluorescent light source selects four specifications identical and the daylight lamp bulb opened simultaneously or close.
Described cell clone survival rate automatic detection device, plank selected by described light barrier, or selects cardboard, or selects plastic plate.
Described cell clone survival rate automatic detection device, described message handler also comprises GPRS data transport module, the input end of described GPRS data transport module is connected with the output terminal of information processing single-chip microcomputer, and the output terminal of described GPRS data transport module is connected with the input end of remote server with internet by GPRS network; Described function button comprises startup/exit button, spectrum arranges button, auto-focusing button, image information collecting button, image noise reduction button, clone's yardstick threshold value arranges button, clone excision and recognition button, clonal survival quantizes button, file save button and GPRS data transmission button.
Described cell clone survival rate automatic detection device, the single-chip microcomputer based on ARM9 kernel selected by described information processing single-chip microcomputer.
A detection method for cell clone survival rate automatic detection device, comprises the following steps:
(1) cell clone sample to be measured and compared with control cells clone sample are placed on respectively in two grooves on the push-and-pull pallet of sample image Acquisition Instrument;
(2) focal length of information processing single-chip microcomputer to ccd video camera adjusts automatically;
(3) information processing Single-chip Controlling three primary light source produces the light of white and default detection spectrum two kinds of colors successively, controls ccd video camera simultaneously and under the reflected light illumination of two kinds of colors, respectively gathers a width cell clone sample image respectively; Information processing Single-chip Controlling three primary light source is closed, and controls fluorescent light source and opens, and controls ccd video camera simultaneously under transmitted light illumination, gathers a width cell clone sample image;
(4) the three width cell clone sample images gathered are sent to information processing single-chip microcomputer by ccd video camera, and information processing single-chip microcomputer is presented in LCD display;
(5) the three width cell clone sample images received are carried out noise reduction process by information processing single-chip microcomputer successively, and in LCD display, show the cell clone sample image after noise reduction;
(6) information processing single-chip microcomputer is according to the cell clone yardstick threshold value preset, and carries out cell clone identification successively, and in LCD display, show the cell clone sample image with clonal marker to three width cell clone sample images after noise reduction;
(7) information processing single-chip microcomputer is according to cell clone recognition result, carries out quantification treatment to the cell clone survival rate of cell clone sample to be measured, and in LCD display, show its cell clone survival rate testing result;
(8) three width cell clone sample images of ccd video camera collection and the cell clone survival rate testing result of cell clone sample to be measured are saved in SD card memory by information processing single-chip microcomputer.
The detection method of described cell clone survival rate automatic detection device, also comprises: three width cell clone sample images of ccd video camera collection and the cell clone survival rate testing result of cell clone sample to be measured are transferred to the remote server on internet by information processing single-chip microcomputer by GPRS data transport module.
The structure of apparatus of the present invention is simple, easy to carry, easily operate, information processing in real time, fast, adopt simultaneously and carry out digital cell clone image information collecting based on multispectral reflected light and transmitted light, relative to prior art, improve the accuracy of detection of cell clone survival rate.Because the pollutant monitoring such as water, soil needs to carry out in the wild usually, the present invention will greatly facilitate the detection of the environmental pollution such as water, soil and monitor in real time, the real-time monitoring of environmental pollution such as air, water, soil and the qualification of each pollutant biology toxicity and evaluation can be applied to, active intervention and the day by day serious problem of environmental pollution of effective suppression are had important practical significance, also cell toxicology scientific research can be applied to, there is provided infrastest data for understanding cell toxicology mechanism, there is important scientific meaning.
Accompanying drawing explanation
Fig. 1 is apparatus structure schematic diagram of the present invention;
Fig. 2 is principle schematic of the present invention;
Fig. 3 is the example images figure of cell clone sample under reflected light and transmitted light.
Embodiment
The present invention is further illustrated below in conjunction with the drawings and specific embodiments.
As shown in Figure 1 and Figure 2, a kind of cell clone survival rate automatic detection device, comprise for the sample image Acquisition Instrument 1 of cell clone sample image digital information acquisition and the message handler 2 for the treatment of cell clone sample image information, sample image Acquisition Instrument 1 is connected with message handler 2 by USB data line.
Sample image Acquisition Instrument 1 comprises rectangular box 10, and the surrounding of rectangular box 10 is made up of the light barrier of white, and the material of light barrier can be plank, or is cardboard, or is plastic plate.High definition ccd video camera 11 for gathering cell clone sample image information is embedded in directly over the top of rectangular box 10, in order to prevent, uneven illumination is even causes information acquisition imperfect, its surrounding has inlayed the three primary light source for providing variable color light, and three primary light source comprises electronic color-changing control circuit board 121 triphosphor tube light 122 identical with four specifications.
In the centre position of rectangular box 10, namely be a L-type push-and-pull clear glass pallet 14 immediately below high definition ccd video camera 11, L-type push-and-pull clear glass pallet 14 is fixed on the centre position of rectangular box 10 by grooved tracks 17 horizontal slip on the inwall of rectangular box 10 both sides.Etch two circular grooves 15,16 in the middle part of L-type push-and-pull clear glass pallet 14, be respectively used to place cell clone sample to be measured and compared with control cells clone sample.Be provided with groove type handle 18 in the centre position, front end of L-type push-and-pull clear glass pallet 14, convenient operation person pulls L-type push-and-pull clear glass pallet 14.
When gathering cell clone sample image information, operator holds the groove type handle 18 outside L-type push-and-pull clear glass pallet 14, grooved tracks 17 horizontal slip of L-type push-and-pull clear glass pallet 14 along both sides is pulled out, cell clone sample to be measured is placed in circular groove 15, compared with control cells clone sample is placed in circular groove 16, then, L-type push-and-pull clear glass pallet 14 back in rectangular box 10 by the grooved tracks 17 then along both sides.
The bottom of rectangular box 10 is inlaid with four daylight lamp bulbs 13, specification identical and simultaneously open or close, for generation of the light of transmission cell clone sample, so that high definition ccd video camera 11 gathers cell clone sample information in transmitted light.
Message handler 2 is " trapezoidal " casing, is fixed on the top of the rectangular box 10 of sample image Acquisition Instrument 1.Message handler 2 comprises that information processing single-chip microcomputer 20, LCD display 21, SD interface 25, GPRS data transport module 23 and 10 function buttons, 24,10 function buttons 24 are startup/exit button 240 respectively, spectrum arranges button 241, auto-focusing button 242, image information collecting button 243, image noise reduction button 244, clone's yardstick threshold value arranges button 245, clone excision and recognition button 246, clonal survival quantizes button 247, file save button 248 and GPRS data transmission button 249.
Information processing single-chip microcomputer 20 is the single-chip microcomputers based on ARM9 kernel, as ARM920TS3C2440.The output terminal of function button 24 is connected with the input end of information processing single-chip microcomputer 20, the output terminal of information processing single-chip microcomputer 20 is connected with the input end of electronic color-changing control circuit board 121, daylight lamp bulb 13, LCD display 21 and GPRS data transport module 23 respectively, information processing single-chip microcomputer 20 is connected with high definition ccd video camera 11 interactive mode, the output terminal of information processing single-chip microcomputer 20 is connected with the input end of SD card memory 22 by SD interface 25, and the output terminal of GPRS data transport module 23 is connected with the input end of internet with remote server 3 by GPRS network.
When triphosphor tube light 122 works, electronic color-changing control circuit board 121 controls the luminosity change of triphosphor tube light 122 according to the principle of three primary colours, realize the automatic change of triphosphor tube light 122 glow color, above different colors and the light-illuminating of spectral band to cell clone sample, the different coloring degree of cell clone sample is made to have different reflecting effects.The multiple spectrum cell clone sample image of collection is read in message handler 2 by sample image Acquisition Instrument 1 automatically, the computer program entrance that each function button 24 on message handler 2 is corresponding different respectively, trigger that different function buttons 24 can show the cell clone sample image gathered, process, quantization cell clonal survival, and data are filed, stored in remote data base.
A detection method for cell clone survival rate automatic detection device, comprises the following steps:
Cell clone sample to be measured and compared with control cells clone sample is placed respectively in S0, two circular grooves 15,16 on the L-type push-and-pull clear glass pallet 14 of sample image Acquisition Instrument 1; Power-on switch 26, power light 27 is bright;
Startup/exit button 240 on S1, trigger message processor 2 panel, active cell clone sample image information collecting and cell clone survival rate automatic checkout system program;
S2, trigger spectrum button 241 is set, select to arrange that to detect spectrum T(can be one in red, green, blue three primary colours, or mixed spectra);
S3, trigger auto-focusing button 242, the focal length of adjustment high definition ccd video camera 11 automatically, make cell clone sample can blur-free imaging, and show image in LCD display 21;
S4, trigger image information acquisition button 243, triphosphor tube light 122 produces the light of white and spectrum T two kinds of colors successively, and high definition ccd video camera 11 is automatically each under the light of different colours simultaneously gathers a width cell clone sample image; Then, triphosphor tube light 122 is closed, four the daylight lamp bulbs 13 in bottom are bright, high definition ccd video camera 11 is captured in the cell clone sample image information under transmitted light illumination, the information at every turn collected all shows in LCD display 21, and after information acquisition, four, bottom daylight lamp bulb 13 is closed, four, top triphosphor tube light 122 is bright, waits for information acquisition next time;
The three width cell clone sample images collected are carried out noise reduction process, the cell clone sample image after LCD display 21 shows noise reduction by S5, trigger image noise reduction button 244 successively;
S6, triggering clone yardstick threshold value arrange button 245, as required, increase or reduce cell clone yardstick threshold value;
Three width cell clone sample images after noise reduction are carried out cell clone identification, and in LCD display 21, show the cell clone sample image with clonal marker by S7, triggering clone excision and recognition button 246 successively;
S8, trigger clonal survival quantize button 247, quantification treatment is carried out to the cell clone survival rate of cell clone sample to be measured, and in LCD display 21 showed cell clonal survival testing result;
S9, triggering filing save button 248, the three width cell clone sample images collected are carried out mark filing, the cell clone survival rate together with the cell clone sample to be measured detected is saved in the database of SD card memory 22 correspondence;
If S10 user needs GPRS network to send information, then trigger GPRS data transmission button 249, the three width cell clone sample image information collected and testing result are transferred to the remote server 3 on internet by GPRS data transport module 23, facilitate user to access and recall;
If S11 needs end program, then trigger startup/exit button 240, exit the information acquisition of cell clone sample image and cell clone survival rate automatic checkout system program.
Fig. 3 (a) is the cell clone sample image that high definition ccd video camera 11 obtains under triphosphor tube light 122 produces white light conditioned reflex, and Fig. 3 (b) is the cell clone sample image that high definition ccd video camera 11 obtains under daylight lamp bulb 13 Transmission Conditions.Can clearly find out from two width images, under transmitted light condition, the cell clone group be much sticked together is divided to come, and this is that the accuracy of detection improving cell clone survival rate is laid a good foundation.
From the above, the present invention gathers the reflection and transmission image information of the cell clone sample under multiple spectrum, using forestland recognition technology carries out the detection of cell clone survival rate.The present invention synchronously reads cell clone sample to be measured and compared with control cells clone sample image by sample image Acquisition Instrument 1, then message handler 2 is sent to, image procossing and recognizer is called respectively in message handler 2, utilize image procossing and the mode identification method of sequencing, determine the cell clone survival rate of cell clone sample to be measured, and the cell clone sample image collected and cell clone survival rate testing result are saved in the database of SD card memory 22 correspondence automatically, subsequently, can also select as required whether by internet transmission on remote server 3.The mode that the present invention adopts cell clone sample to be measured and compared with control cells clone sample image information to gather simultaneously, effectively prevent the cell clone sample image process caused by the difference of illumination background and identification error.
Present invention achieves cell clone sample to be measured and compared with control cells clone sample image synchronization, automation collection, Digital Image Processing and mode identification technology is adopted to carry out processing to the cell clone sample image information collected, wherein, message handler by information acquisition, image display, noise reduction and identification, clonal survival quantitatively, data filing storage, the function i ntegration such as GPRS data transmission is in integrally.Relative to the artificial quantivative approach of existing cell clone survival rate, the present invention is by automatic acquisition and processing cell clone sample image information, computerize is carried out to cell clone survival rate quantitative, simple to operate, convenient, avoid the metrical error caused by manual type, substantially increase accuracy of detection, efficiency, save human cost, meanwhile, Detection Information filing and hold function facilitate the return visit of testing process and review.
The above embodiment is only be described the preferred embodiment of the present invention; not scope of the present invention is limited; under not departing from the present invention and designing the prerequisite of spirit; the various distortion that those of ordinary skill in the art make technical scheme of the present invention and improvement, all should fall in protection domain that claims of the present invention determine.
Claims (9)
1. a cell clone survival rate automatic detection device, it is characterized in that: described cell clone survival rate automatic detection device comprises sample image Acquisition Instrument and message handler, the push-and-pull pallet that described sample image Acquisition Instrument comprises rectangular box, is arranged on the ccd video camera of rectangular box inside top central authorities, is arranged on the three primary light source of rectangular box inside top surrounding, is arranged on the fluorescent light source of rectangular box bottom inside central authorities and is arranged in rectangular box; The sidepiece of described rectangular box adopts light barrier, and the central part of described push-and-pull pallet offers two grooves, is respectively used to place cell clone sample to be measured and compared with control cells clone sample;
Described message handler is connected with sample image Acquisition Instrument by USB data line, described message handler comprises information processing single-chip microcomputer, LCD display, SD interface and some function buttons, the output terminal of described function button is connected with the input end of information processing single-chip microcomputer, the output terminal of described information processing single-chip microcomputer respectively with three primary light source, fluorescent light source is connected with the input end of LCD display, described information processing single-chip microcomputer is connected with ccd video camera interactive mode, the output terminal of described information processing single-chip microcomputer is connected with the input end of SD card memory by SD interface.
2. cell clone survival rate automatic detection device according to claim 1, is characterized in that: the both sides inwall of described rectangular box is respectively equipped with the grooved tracks for placing push-and-pull pallet.
3. cell clone survival rate automatic detection device according to claim 1, it is characterized in that: the cross section of described push-and-pull pallet is L-type, its long leg is horizontally set in rectangular box, its short leg is vertically arranged on rectangular box front end, and the central part of described short leg is provided with groove type handle.
4. cell clone survival rate automatic detection device according to claim 1, it is characterized in that: described three primary light source comprises the electronic color-changing control circuit board triphosphor tube light identical with four specifications, the input end of described electronic color-changing control circuit board is connected with the output terminal of information processing single-chip microcomputer, and the described output terminal of electronic color-changing control circuit board is connected with the input end of triphosphor tube light; Described fluorescent light source selects four specifications identical and the daylight lamp bulb opened simultaneously or close.
5. cell clone survival rate automatic detection device according to claim 1, is characterized in that: plank selected by described light barrier, or selects cardboard, or selects plastic plate.
6. cell clone survival rate automatic detection device according to claim 1, it is characterized in that: described message handler also comprises GPRS data transport module, the input end of described GPRS data transport module is connected with the output terminal of information processing single-chip microcomputer, and the output terminal of described GPRS data transport module is connected with the input end of remote server with internet by GPRS network; Described function button comprises startup/exit button, spectrum arranges button, auto-focusing button, image information collecting button, image noise reduction button, clone's yardstick threshold value arranges button, clone excision and recognition button, clonal survival quantizes button, file save button and GPRS data transmission button.
7. cell clone survival rate automatic detection device according to claim 1, is characterized in that: the single-chip microcomputer based on ARM9 kernel selected by described information processing single-chip microcomputer.
8. the detection method of a kind of cell clone survival rate automatic detection device according to claim 1, is characterized in that, comprise the following steps:
(1) cell clone sample to be measured and compared with control cells clone sample are placed on respectively in two grooves on the push-and-pull pallet of sample image Acquisition Instrument;
(2) focal length of information processing single-chip microcomputer to ccd video camera adjusts automatically;
(3) information processing Single-chip Controlling three primary light source produces the light of white and default detection spectrum two kinds of colors successively, controls ccd video camera simultaneously and under the reflected light illumination of two kinds of colors, respectively gathers a width cell clone sample image respectively; Information processing Single-chip Controlling three primary light source is closed, and controls fluorescent light source and opens, and controls ccd video camera simultaneously under transmitted light illumination, gathers a width cell clone sample image;
(4) the three width cell clone sample images gathered are sent to information processing single-chip microcomputer by ccd video camera, and information processing single-chip microcomputer is presented in LCD display;
(5) the three width cell clone sample images received are carried out noise reduction process by information processing single-chip microcomputer successively, and in LCD display, show the cell clone sample image after noise reduction;
(6) information processing single-chip microcomputer is according to the cell clone yardstick threshold value preset, and carries out cell clone identification successively, and in LCD display, show the cell clone sample image with clonal marker to three width cell clone sample images after noise reduction;
(7) information processing single-chip microcomputer is according to cell clone recognition result, carries out quantification treatment to the cell clone survival rate of cell clone sample to be measured, and in LCD display, show its cell clone survival rate testing result;
(8) three width cell clone sample images of ccd video camera collection and the cell clone survival rate testing result of cell clone sample to be measured are saved in SD card memory by information processing single-chip microcomputer.
9. the detection method of cell clone survival rate automatic detection device according to claim 8, it is characterized in that, also comprise: three width cell clone sample images of ccd video camera collection and the cell clone survival rate testing result of cell clone sample to be measured are transferred to the remote server on internet by information processing single-chip microcomputer by GPRS data transport module.
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