CN106841072A - A kind of method differentiated for algae with algae proliferation non-destructive monitoring situation - Google Patents
A kind of method differentiated for algae with algae proliferation non-destructive monitoring situation Download PDFInfo
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- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
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Abstract
The present invention discloses a kind of method differentiated for algae with algae proliferation non-destructive monitoring situation, comprises the following steps：1) using a kind of spectrum micro-image for differentiating for algae and sample being obtained with the system of algae proliferation non-destructive monitoring situation；2) spectrum micro-image is removed into background by binary conversion treatment, and carries out rim detection, so as to obtain the spectral information of sample；3) spectral information extracted, differentiated, and counted various algae numbers；4) sample is repeated the above steps daily, various algae data can be obtained by long-term observation.Instant invention overcomes traditional chemical routes can disturbed specimen and its surrounding enviroment shortcoming, realize Non-Destructive Testing, the algae in water body can be carried out repeatedly with lasting monitoring, additionally it is possible to obtain more and more accurately sample message.
Description
Technical field
Differentiate for algae and algae the present invention relates to spectral imaging technology and spectral unmixing technical field, more particularly to one kind
The method that class breeds non-destructive monitoring situation.
Background technology
Due to reasons such as body eutrophications, " red tide " phenomenon occurs in ocean or fresh water water system." red tide "
Phenomenon refers to the explosive propagation of some phytoplanktons, protozoan or bacterium or height in water body under certain environmental conditions
Assemble and cause one kind of water color-changing to be harmful to ecology phenomenon.The toxin of red tide plankton release toxin red tide plankton release can cause
The biological deaths such as marine fish, shrimp, shellfish, or toxin is finally included the mankind to their other animals that ingest in being enriched in marine product
Toxic action is produced, red tide plankton largely consumes oxygen in addition, causes Dissolved Oxygen in Water to reduce, and can also make marine organisms because of anoxic
And mortality.Used as the key factor that red tide is formed, breeding situation of the algae in water body receives much concern.
However, still lack limited research tool and research method at present being carried out to the breeding situation of various algae in water body
Lasting monitoring.Although observing water body using conventional microscope, Non-Destructive Testing can be realized, and then can continue to monitor,
Measures of Algae in Water Body complicated component, the classification of some algae can not be differentiated only from pattern, it is also difficult to be counted with naked eyes
Measures of Algae in Water Body number.Although and very accurate using the method for chemical labeling and be easy to statistics, algae itself and residing
Environment receives severe jamming, or even directly affects the growth of algae, therefore the also improper algae for continuing to monitor in water body
Breeding situation.
A kind of apparatus and method that lasting accurate measurements can be carried out to Measures of Algae in Water Body breeding situation are urgently developed at present,
This will be helpful to promote the research related to red tide, for the generation for preventing and controlling red tide provides strong support.In view of various
The respective pigment composition of algae and component have difference, therefore its color or autofluorescence also have difference, can be by light
Spectrum information carries out species discriminating to algae.
Therefore, the prior art is defective, it is necessary to improve.
The content of the invention
It is a primary object of the present invention to overcome it is a primary object of the present invention to overcome traditional detection means and detection side
Method cannot meet carries out the shortcoming of accurate and lossless monitoring to Measures of Algae in Water Body breeding situation, propose a kind of spectral unmixing technology and
The brand-new monitoring method that spectral imaging technology is combined, is devoted to promoting the development of red tide correlative study.
The invention discloses one kind for continuing interference-free monitoring Complex water body algae in environment species and proliferative conditions
Method.By carrying out microspectrum imaging to the algae in water body, and carried out by obtaining the spectral information of each of which algae
Compare, so as to differentiate to algae classification, and counted.
Laser system of the present invention is realized by following technical scheme：
A kind of method differentiated for algae with algae proliferation non-destructive monitoring situation, comprises the following steps：
1) differentiate micro- with the spectrum of the system acquisition sample of algae proliferation non-destructive monitoring situation for algae using a kind of
Image；
2) spectrum micro-image is removed into background by binary conversion treatment, and carries out rim detection, so as to obtain sample
Spectral information；
3) spectral information extracted, differentiated, and counted various algae numbers；
4) sample is repeated the above steps daily, various algae data can be obtained by long-term observation.
Further, described discriminating for algae includes with the system of algae proliferation non-destructive monitoring situation：Lighting source, load
Thing platform, microcobjective, two are to dichronic mirror, the one 45 ° of speculum, 45 ° of semi-transparent semi-reflecting lens, the 2nd 45 ° of speculum, liquid crystal filters
Part, the first imaging len, the second imaging len, Grey imaging device, color imaging device, excite photosystem and collimation lens
Group；
It is described to excite the light that photosystem is launched to be penetrated two on dichronic mirror through collimation lens set, then by two to dichronic mirror
Reflection is injected in microscope from the bottom to top；The light projection that lighting source is projected on objective table, then through the sample on objective table,
Project into microscope from top to bottom, and be incident upon on the one 45 ° of speculum to dichronic mirror through two.One 45 ° of speculum is again
Light is reflexed into 45 ° of semi-transparent semi-reflecting lens.Reflex to the light on 45 ° of semi-transparent semi-reflecting lens and be divided into two-way, light is by 45 ° semi-transparent half all the way
Injected in color imaging device through the first imaging len after anti-mirror reflection；Another road light after 45 ° of semi-transparent semi-reflecting lens through penetrating
On 2nd 45 ° of speculum, after the 2nd 45 ° of light of speculum reflection is through liquid crystal filtering device and the second imaging len, most
Eventually in input Grey imaging device.
Further, the liquid crystal filter part is filtered regulation to visible light wave range (380-760nm)；Described swashs
Luminescent system is short wavelength's (wavelength<410nm) light source, the described light source for exciting photosystem is ultraviolet lamp, ultraviolet LD and
Ultraviolet LED；The one 45 ° of speculum, 45 ° of semi-transparent semi-reflecting lens, the wave band of the light of the 2nd 45 ° of speculum reflection are visible light wave
Section；The spectral resolution of described liquid crystal filter reaches 1nm, can reflect into the flashlight for coming to the second speculum and filter
Ripple, thus export Single wavelength flashlight and by the second imaging len in Grey imaging device into the gray-scale map under Single wavelength
Picture.
Further, the method for the spectrum micro-image of the use image device acquisition sample is comprised the following steps：
1) the algae sample of the water content that will be placed in transparent culture dish is placed on the objective table of system, opens illumination light
Source, liquid crystal filter part and image device, the light that lighting source sends are formed and carry algae sample by being partly absorbed after sample
This flashlight；
2) regulation microcobjective position is until it is observed that clearly in Grey imaging device and color imaging device
Imaging, so as to obtain transmitted spectrum imaging；
3) liquid crystal filter part filter range and scanning accuracy are set, the image device time for exposure is set, unlatching is continuously swept
Screening-mode is retouched, a series of spectrum micro-images under one section of optical wavelength interval are obtained, the spectrum micro-image is transmitted light
Spectrum imaging.
Further, the method for the spectrum micro-image of the use image device acquisition sample is comprised the following steps：
1) the algae sample of the water content that will be placed in transparent culture dish is placed on the objective table of system, opens exciting light
System, liquid crystal filter part and image device, the light that lighting source sends are formed and carry algae by being partly absorbed after sample
The flashlight of sample；
2) regulation microcobjective position is until it is observed that clearly in Grey imaging device and color imaging device
Imaging, so as to obtain transmitted spectrum imaging；
3) liquid crystal filter part filter range and scanning accuracy are set, the image device time for exposure is set, unlatching is continuously swept
Screening-mode is retouched, a series of spectrum micro-images under one section of optical wavelength interval are obtained, the spectrum micro-image is fluorescence light
Spectrum imaging.
Further, it is described that spectrum micro-image is comprised the following steps by the method that binary conversion treatment removes background：
1) grey value histograms of entire image are first counted, then original image is divided into prospect, background two by setting threshold value
Individual image；
2) background is set to 0 gray value, sample is set to highest gray value, so as to by measured object and background separation.
Further, described Edge check is used to sketching the contours the profile of each frond in image, and by all pictures in profile
Vegetarian refreshments is classified as the internal information of a certain frond, so that reasonable drawing can be carried out to each frond spectral information, and can be to frond
Number is checked；Described Edge check by based on the image after binaryzation, by the ash between background and measured object
Degree difference carries out edge judgement；The method of the Edge check includes：Method of differential operator, Model match method, Wavelet Detection method, god
Through network technique.
Further, the method for the withdrawing spectral information is comprised the following steps：
1) object pixel point coordinates is extracted, the x-axis and y of the corresponding target pixel points of data spectrum picture is manually selected
Axial coordinate value, or by double mouse click target pixel points, obtain corresponding x-axis and y-axis coordinate value at the point；
2) after obtaining the coordinate value of target pixel points, gray value at the spectrum picture coordinate value is read under each wavelength,
Intensity when the gray value size represents the target pixel points in the wavelength；
3) then after the pixel gray level Value Data of picture under having read all wavelengths, with wavelength value as x-axis, with gray scale
It is worth for y-axis can describe the corresponding curve of spectrum of the pixel；
Further, described algae discrimination method is the spectrum for surveying pixel with target using the curve of spectrum of standard items
Curve ratio compared with；The object pixel point selection is located at the pixel within algae edge.
Further, the statistical method of the algae is after carrying out species discriminating to each frond, in spectrum micro-image
Middle use different colours are marked to different algal species, the pixel being located within algae edge are calculated as into a frond, and mark
Same color is designated as, and counts same color frond number.
Using such scheme, the present invention compared with prior art, has the following advantages that and beneficial effect：
1st, instant invention overcomes traditional chemical routes meeting disturbed specimen and its shortcoming of surrounding enviroment, lossless inspection is realized
Survey, the algae in water body can be carried out repeatedly with lasting monitoring.
2nd, the system and method that the present invention is provided is obtained in that more and more accurately sample message, micro- relative to tradition
Mirror, in addition to it can obtain its topographical information, additionally it is possible to obtain the spectral information of each frond, and can intuitively by various algae
Distribution situation be objectively displayed on picture, realize to the accurate statistics of each algae.Though additionally, traditional chemical method
So can also distinguish frond species and number can be counted, but cannot also obtain topographical information, acquired information content is far low
In the system and method that the present invention is provided.
3rd, the system operatio that the present invention is provided is simple, and detection is accurate quick, and strong applicability is weaker for autofluorescence
Algae can be detected that less for pigment, transparency algae higher can use fluorescence spectrum using transmitted spectrum imaging
Imaging is detected, it is also possible to both approaches are respectively adopted sample is detected, obtains the standard of more information and Geng Gao
True rate.
Brief description of the drawings
Fig. 1 is structural representation of the invention；
Fig. 2 is withdrawing spectral information process schematic of the invention；
Fig. 3 is operational flowchart of the invention；
Fig. 4 is the identification result displaying figure of case study on implementation in the present invention.
Specific embodiment
Below in conjunction with the drawings and specific embodiments, the present invention is described in detail.
With reference to embodiment and accompanying drawing, the present invention is described in further detail, but embodiments of the present invention are not limited
In this.
Referring to figs. 1 to shown in Fig. 4, in the present invention for continue interference-free monitoring Complex water body algae in environment species and
The microscopic system of proliferative conditions includes lighting source 1, three-dimensional adjustable objective table 2, microcobjective 3, two to the 4, the 1st ° of dichronic mirror
The 6, the 2nd 45 ° of semi-transparent semi-reflecting lens of 5,45 ° of speculum speculum 7, liquid crystal filter part 8, the first imaging len 9, second are imaged saturating
Mirror 10, Grey imaging device 11, color imaging device 12, excite photosystem 13 and collimation lens set 14.
The light that photosystem 13 is launched is excited to be penetrated two on dichronic mirror 4 through collimation lens set 14, then by two to color separation
Mirror 4 reflects and inject from the bottom to top in microscope.The light projection that lighting source 1 is projected is on three-dimensional adjustable objective table 2, then passes through load
Sample on thing platform 2, projects into microscope from top to bottom, and is incident upon on the one 45 ° of speculum 5 to dichronic mirror 4 through two.
Light is reflexed to 45 ° of semi-transparent semi-reflecting lens 6 by the one 45 ° of speculum 5 again.The light reflexed on 45 ° of semi-transparent semi-reflecting lens 6 is divided into two
Road, injects in color imaging device 12 after light is reflected by 45 ° of semi-transparent semi-reflecting lens 6 all the way through the first imaging len 9；Another road light
Through being penetrated on the 2nd 45 ° of speculum 7 after 45 ° of semi-transparent semi-reflecting lens 6, the light reflected by the 2nd 45 ° of speculum 7 is filtered through liquid crystal
After the imaging len 10 of wave device 8 and second, in final input Grey imaging device 11.
Wherein, the described photosystem 13 that excites is short wavelength's (wavelength<410nm) light source, the described photosystem 13 that excites
Light source is ultraviolet lamp, ultraviolet LD and ultraviolet LED；The wave band of the light of the speculum reflection is visible light wave range.Described
Collimation lens set 14 has three convex lens groups into exciting photosystem 13 to be located at front lens focal point, front lens and intermediate lens
Distance do not limit, intermediate lens and rear lenses focal length are f2And f3, then both distances are f2+f3, exciting light put by the system
Big multiple is A=f2/f3。
The lighting source 1 can be the broad spectrum light sources such as high pressure xenon lamp or Halogen lamp LED, in order to be used as transmitted light
Spectrum imaging.The microcobjective 3 needs, using achromatism infinity system microcobjective 3, to be easy to add in the optical path or move
Imaging surface of the eliminating optical element without change subsequent optical path.It is described excite photosystem 13 should be short wavelength blue violet light or
Ultraviolet light, such as ultraviolet laser, ultraviolet LED or uviol lamp etc..Described two need to use to being shorter than excitation wavelength to dichronic mirror 4
Light is reflected, and the light for being longer than excitation wavelength is transmitted, the wavelength of the autofluorescence launched by sample than
The optical wavelength for exciting photosystem 13 to be launched will be grown, therefore two can completely cut off exciting light into subsequent optical path system to dichronic mirror 4
System, interference is produced into fluorescence signal.
Because transmitted spectrum of the present invention imaging and fluorescence spectrum imaging are all the liquid crystal filters in visible-range
Wave device 8 should be selected and be filtered regulation to visible light wave range (400nm-700nm).The spectrum of described liquid crystal filter point
Resolution reaches 1nm, can reflect into the flashlight for coming to the second speculum and be filtered, so as to export Single wavelength flashlight and pass through
The second imaging len 10 is crossed in Grey imaging device 11 into the gray level image under Single wavelength.Need to set one simultaneously in system
Grey imaging device 11 coordinates liquid crystal filter to realize light spectrum image-forming, and a colour for gray scale CCD or COMS camera
Image device 12 --- colored CCD or COMS cameras, to obtain the imaging under sample true colors.
Differentiate the method with algae proliferation non-destructive monitoring situation for algae, comprise the following steps：
1) differentiate micro- with the spectrum of the system acquisition sample of algae proliferation non-destructive monitoring situation for algae using a kind of
Image；
2) spectrum micro-image is removed into background by binary conversion treatment, and carries out rim detection, so as to obtain sample
Spectral information；
3) spectral information extracted, differentiated, and counted algae number.
Wherein, differentiate that can obtain two kinds of spectrum with the system of algae proliferation non-destructive monitoring situation shows for algae using a kind of
Micro- image, one kind is transmitted spectrum imaging, and another kind is fluorescence spectrum imaging.
The method that the use image device obtains the spectrum micro-image of sample transmitted spectrum imaging is comprised the following steps：
1) the algae sample of the water content that will be placed in transparent culture dish is placed on the objective table 2 of system, opens illumination
Light source 1, liquid crystal filter part 8 and image device, by being partly absorbed after sample, formation is carried the light that lighting source 1 sends
The flashlight of algae sample；
2) the regulation position of microcobjective 3 is until it is observed that clear in Grey imaging device 11 and color imaging device 12
Clear imaging, so as to obtain transmitted spectrum imaging；
3) filter range of liquid crystal filter part 8 and scanning accuracy are set, the image device time for exposure is set, unlatching is continuously swept
Screening-mode is retouched, a series of spectrum micro-images under one section of optical wavelength interval are obtained, the spectrum micro-image is transmitted light
Spectrum imaging.
When closing excites photosystem 13, only open white lumination system, the illumination light by after sample, being partly absorbed,
Form the flashlight with absorption information.This flashlight is collected by microcobjective 3, by two to dichronic mirror 4, speculum and semi-transparent
It is divided into two-way after semi-reflective mirror, forms coloured image in color imaging device 12 by imaging len all the way.Another road signal
Light is then reflected by a reflector, into liquid crystal filter.Liquid crystal filter is scanned by the spectral resolution with 1nm, every turn
Change a filter wavelength, carried out by imaging len and Grey imaging device 11 it is Polaroid, until scanning whole visible light wave
Section, obtains the gray level image under each wavelength, i.e. transmitted spectrum imaging.
The method that the use image device obtains the spectrum micro-image of sample fluorescence light spectrum image-forming is comprised the following steps：
1) the algae sample of the water content that will be placed in transparent culture dish is placed on the objective table 2 of system, and unlatching is excited
Photosystem 13, liquid crystal filter part 8 and image device, the light that lighting source 1 sends form band by being partly absorbed after sample
There is the flashlight of algae sample；
2) the regulation position of microcobjective 3 is until it is observed that clear in Grey imaging device 11 and color imaging device 12
Clear imaging, so as to obtain transmitted spectrum imaging；
3) filter range of liquid crystal filter part 8 and scanning accuracy are set, the image device time for exposure is set, unlatching is continuously swept
Screening-mode is retouched, a series of spectrum micro-images under one section of optical wavelength interval are obtained, the spectrum micro-image is fluorescence light
Spectrum imaging.
When white lumination system is closed, only open and excite the photosystem 13, exciting light will to reflex to load to dichronic mirror 4 by two
Sample on thing platform 2, and the fluorescence of its own is excited, fluorescence signal is collected by microcobjective 3, by two to dichronic mirror 4, reflection
It is divided into two-way after mirror and semi-transparent semi-reflecting lens, forms coloured image in color imaging device 12 by imaging len all the way.Separately
Flashlight is then reflected by a reflector all the way, into liquid crystal filter.Liquid crystal filter is swept by the spectral resolution with 1nm
Retouch, often change a filter wavelength, carried out by imaging len and Grey imaging device 11 it is Polaroid, until scanning entirely may be used
See optical band, obtain the gray level image under each wavelength, i.e. fluorescence spectrum imaging.
After sample spectra micro-image is obtained, in addition it is also necessary to by background removal, Edge check, withdrawing spectral information, light
After the steps such as spectrum information discriminating, quantity statistics, discriminating and the statistics of algae number of algal kind are finally realized.
Specifically, background removal be in order to obtain spectrum micro-imaging after algae and background are separated, accordingly, it would be desirable to
Process image binaryzation and carry out Edge check.Wherein, binaryzation can be realized using OTSU algorithms.It is described by spectrum micrograph
As the method that background is removed by binary conversion treatment is comprised the following steps：
1) grey value histograms of view picture spectrum micro-image are first counted, then before original image is divided into by setting threshold value
Scape, two images of background；
2) background is set to 0 gray value, sample is set to highest gray value, so as to by measured object and background separation.
Described Edge check is used to sketch the contours the profile of each frond in image, and all pixels point in profile is classified as into certain
The internal information of one frond, so as to reasonable drawing can be carried out to each frond spectral information, and can check frond number；
Described Edge check is based on the image after binaryzation, side is carried out by the gray difference between background and measured object
Along judgement；The method of the Edge check includes：Method of differential operator, Model match method, Wavelet Detection method, neural network.
The pixel gray value of spectrum picture captured under a certain optical wavelength is represented into light intensity value, you can represent the picture
Vegetarian refreshments transmission at that wavelength or fluorescence intensity.Once pixel point selection is inside certain frond, it is possible to know this
Frond transmission at that wavelength or fluorescence intensity.Describe wavelength with intensity by scanning the spectrum picture under all wavelengths
The curve of spectrum, you can complete withdrawing spectral information.The method of the withdrawing spectral information is comprised the following steps：
1) object pixel point coordinates is extracted, the x-axis and y of the corresponding target pixel points of data spectrum picture is manually selected
Axial coordinate value, or by double mouse click target pixel points, obtain corresponding x-axis and y-axis coordinate value at the point；
2) after obtaining the coordinate value of target pixel points, gray value at the spectrum picture coordinate value is read under each wavelength,
Intensity when the gray value size represents the target pixel points in the wavelength；
3) then after the pixel gray level Value Data of picture under having read all wavelengths, with wavelength value as x-axis, with gray scale
It is worth for y-axis can describe the corresponding curve of spectrum of the pixel；
Described algae discrimination method is that the curve of spectrum for surveying pixel with target using the curve of spectrum of standard items compares；
The object pixel point selection is located at the pixel within algae edge.
The statistical method of the algae is after carrying out species discriminating to each frond, using difference in spectrum micro-image
Color is marked to different algal species, and the pixel being located within algae edge is calculated as into a frond, and labeled as same
Color, and count same color frond number.
Spectral information can be differentiated and then to algae using methods such as ratio method, neural network algorithm or genetic algorithms
Class is differentiated.After the classification for identifying each frond, according to Edge check result, with reference to frond classification, to various classifications
Algae carry out quantity statistics, under long-term monitoring become can study breeding situation of the various algae in water body.
Embodiment 1
As a kind of case study on implementation, using chlorella and Nannochloropsis oculata as monitor sample in the present invention, in culture dish
Cultivated in water body.Both algae patterns are similar, and volume is tiny, it is difficult to differentiated using conventional microscope, and two
Fluorescent material is less contained by person, therefore the implementation case is tested using transmittance spectrum imaging mode.
The selected lighting source 1 of the implementation case is high pressure xenon lamp, the VariSpecTM type liquid produced using CRi companies
Brilliant wave filter 400-720nm is filtered.Additionally, selection is high to below 400nm wavelength anti-(99.2% reflectivity) and right
400nm wavelengths above is anti-reflection (99.8% transmitance) two are to dichronic mirror 4.Grey imaging device 11 and color imaging device 12
Gray scale CCD and colored CCD are selected respectively.
The semi-transparent semi-reflecting lens selected in the implementation case are semi-transparent semi-reflecting to whole visible light wave range (380nm~780nm),
One 45 ° of speculum of selection and the 2nd 45 ° of speculum are anti-(99.8% reflectivity) high to whole visible light wave range.
In order to obtain larger observation scope, selection multiplication factor is 20 times of infinity system achromatism to the implementation case
Microcobjective 3.
The sweep limits that the implementation case sets liquid crystal filter part 8 is 450nm~700nm, and scanning accuracy is 2nm.If
It is 0.5ms to put the colored CCD time for exposure, and the gray scale CCD time for exposure is 3ms.
The implementation case carries out binary conversion treatment using spectrum picture and OTSU algorithms captured under 550nm wavelength,
Edge check is carried out using Model match method and is sketched the contours, adopted and spectral information is differentiated with ratio method.
The operating mechanism of the system is as follows：After opening lighting source 1, the illumination light in broadband can be through sample by micro- thing
Mirror 3 is collected.During through sample, because algae itself contains various pigments, partial illumination light can be absorbed, thoroughly
The light for shooting out than enter sample before light have compared with lossy, the light without only being passed through from water body by sample be then lost compared with
It is few.Different algae has different absorption losss to the light of different wave length, and these have just taken information by the light after absorption loss
Amount, i.e. its spectral information.
Light is reflected by 45 ° of speculums to dichronic mirror 4 by microcobjective 3 and two and respectively enters two by semi-transparent semi-reflecting lens
Individual imaging optical path, directly carries out colour imaging by imaging len all the way, leads up to liquid crystal filter part 8 and filters, and only allows certain
One wavelength light enters imaging len, and is finally imaged in gray scale CCD.What is shot each time due to gray scale CCD is all certain
Picture under one wavelength, then the gray value of each pixel represents the intensity that the position shows at that wavelength on its picture.It is false
If without algae background gray levels being 200 in the spectrum picture of certain wavelength, the pixel corresponding grey scale value on certain sample is 100, then
Absorptivity [(200-100)/200] the * 100%=50% of the pixel to the wavelength can be calculated, by counting the pixel
Absorptivity or transmitance under all scanning wavelengths of point can then depict the absorption corresponding to the material representated by this pair of point
Or transmission spectrum curve.Because different algal species have different optical absorption characteristics, therefore by absorption curve or transmission curve
Its species can then be differentiated, and finally count its quantity respectively.
The implementation case can obtain identification result image as described in Figure 4 according to flow described in above-mentioned setting and Fig. 3,
It is marked with two pseudo- gray scale fronds different to two classes respectively in the implementation case, is can see in left figure, from pattern
Both cannot be differentiated, but by both spectrum of contrast, can be differentiated, as a result as shown at right.After identification result is obtained
The number of various algae in water body can be counted.
In sum, the present invention first combines spectral imaging technology with algae discriminating, according to the tiny needs of algae
Microscopic system is imaged, and need by spectrum carry out Undamaged determination and count requirement, it is proposed that a kind of spectrum it is micro- into
As system, and obtain data for the system and propose using a kind of realization of algorithm, the algae compared based on spectral information
The method that species differentiates and various algae numbers are counted.
Algae reproduction situation is observed relative to using conventional microscope, system of the present invention is except that can obtain shape
Outside looks information, moreover it is possible to obtain more objective spectral information.Additionally, the present invention is carried out using algorithm to the algae in range of observation
Accurate number statistics, than being differentiated more accurate by micro- sem observation and then artificial counting or chemical method.Pass through
The system and correlation method, can obtain algae micro-imaging, light spectrum image-forming, the various algae curves of spectrum and various algae
The abundant information content such as number and distribution situation.
Above-described embodiment is the present invention preferably implementation method, but embodiments of the present invention are not by above-described embodiment
Limitation, it is other it is any without departing from Spirit Essence of the invention and the change, modification, replacement made under principle, combine, simplification,
Equivalent substitute mode is should be, is included within protection scope of the present invention.
Claims (10)
1. it is a kind of for algae differentiate and algae proliferation non-destructive monitoring situation method, it is characterised in that comprise the following steps：
1) using a kind of spectrum micro-image for differentiating for algae and sample being obtained with the system of algae proliferation non-destructive monitoring situation；
2) spectrum micro-image is removed into background by binary conversion treatment, and carries out rim detection, so as to obtain the spectrum of sample
Information；
3) spectral information extracted, differentiated, and counted various algae numbers；
4) sample is repeated the above steps daily, various algae data can be obtained by long-term observation.
2. it is according to claim 1 for algae differentiate and algae proliferation non-destructive monitoring situation method, it is characterised in that
Described discriminating for algae includes with the system of algae proliferation non-destructive monitoring situation：Lighting source, objective table, microcobjective, two
To dichronic mirror, the one 45 ° of speculum, 45 ° of semi-transparent semi-reflecting lens, the 2nd 45 ° of speculum, liquid crystal filter part, the first imaging len,
Second imaging len, Grey imaging device, color imaging device, excite photosystem and collimation lens set；
It is described to excite the light that photosystem is launched to be penetrated two on dichronic mirror through collimation lens set, then by two to dichroic mirror
In injecting microscope from the bottom to top；The light projection that lighting source is projected on objective table, then through the sample on objective table, from upper
Projected into microscope under, and be incident upon on the one 45 ° of speculum to dichronic mirror through two, the one 45 ° of speculum is again by light
Line reflection is to 45 ° of semi-transparent semi-reflecting lens；Reflex to the light on 45 ° of semi-transparent semi-reflecting lens and be divided into two-way, light is by 45 ° of semi-transparent semi-reflecting lens all the way
Injected in color imaging device through the first imaging len after reflection；Another road light after 45 ° of semi-transparent semi-reflecting lens through penetrating second
It is final to throw after the 2nd 45 ° of light of speculum reflection is through liquid crystal filtering device and the second imaging len on 45 ° of speculums
Enter in gray scale image device.
3. it is according to claim 2 for algae differentiate and algae proliferation non-destructive monitoring situation system, it is characterised in that
The liquid crystal filter part is filtered regulation to visible light wave range；Described excites photosystem for short wavelength light source, described
The light source for exciting photosystem is ultraviolet lamp, ultraviolet LD and ultraviolet LED；The one 45 ° of speculum, 45 ° of semi-transparent semi-reflecting lens,
The wave band of the 2nd 45 ° of light of speculum reflection is visible light wave range；The spectral resolution of described liquid crystal filter reaches 1nm,
The flashlight for coming can be reflected into the second speculum to be filtered, so as to export Single wavelength flashlight and be imaged by second saturating
Mirror is in Grey imaging device into the gray level image under Single wavelength.
4. it is according to claim 3 for algae differentiate and algae proliferation non-destructive monitoring situation method, it is characterised in that
The method that the use image device obtains the spectrum micro-image of sample is comprised the following steps：
1) the algae sample of the water content that will be placed in transparent culture dish is placed on the objective table of system, unlatching lighting source,
Liquid crystal filter part and image device, the light that lighting source sends are formed and carry algae sample by being partly absorbed after sample
Flashlight；
2) regulation microcobjective position is until it is observed that be clearly imaged in Grey imaging device and color imaging device,
So as to obtain transmitted spectrum imaging；
3) liquid crystal filter part filter range and scanning accuracy are set, the image device time for exposure is set, open continuous scanning and clap
Take the photograph pattern, obtain a series of interval lower spectrum micro-images of one section of optical wavelength, the spectrum micro-image for transmitted spectrum into
Picture.
5. it is according to claim 3 for algae differentiate and algae proliferation non-destructive monitoring situation method, it is characterised in that
The method that the use image device obtains the spectrum micro-image of sample is comprised the following steps：
1) the algae sample of the water content that will be placed in transparent culture dish is placed on the objective table of system, and unlatching excites spectrum
System, liquid crystal filter part and image device, the light that lighting source sends are formed and carry algae sample by being partly absorbed after sample
This flashlight；
2) regulation microcobjective position is until it is observed that be clearly imaged in Grey imaging device and color imaging device,
So as to obtain transmitted spectrum imaging；
3) liquid crystal filter part filter range and scanning accuracy are set, the image device time for exposure is set, open continuous scanning and clap
Take the photograph pattern, obtain a series of interval lower spectrum micro-images of one section of optical wavelength, the spectrum micro-image for fluorescence spectrum into
Picture.
6. it is according to claim 1 for algae differentiate and algae proliferation non-destructive monitoring situation method, it is characterised in that
It is described that spectrum micro-image is comprised the following steps by the method that binary conversion treatment removes background：
1) grey value histograms of view picture spectrum micro-image are first counted, then original image is divided into prospect by setting threshold value, carried on the back
Two images of scape；
2) background is set to 0 gray value, sample is set to highest gray value, so as to by measured object and background separation.
7. it is according to claim 1 for algae differentiate and algae proliferation non-destructive monitoring situation method, it is characterised in that
Described Edge check is used to sketch the contours the profile of each frond in image, and all pixels point in profile is classified as into a certain frond
Internal information, so as to reasonable drawing can be carried out to each frond spectral information, and can check frond number；Described side
Along detection based on the image after binaryzation, edge judgement is carried out by the gray difference between background and measured object；
The method of the Edge check includes：Method of differential operator, Model match method, Wavelet Detection method, neural network.
8. it is according to claim 1 for algae differentiate and algae proliferation non-destructive monitoring situation method, it is characterised in that
The method of the withdrawing spectral information is comprised the following steps：
1) object pixel point coordinates is extracted, manually selects the x-axis and y-axis of the corresponding target pixel points of data spectrum picture to sit
Scale value, or by double mouse click target pixel points, obtain corresponding x-axis and y-axis coordinate value at the point；
2) after obtaining the coordinate value of target pixel points, gray value, the ash at the spectrum picture coordinate value are read under each wavelength
Intensity when angle value size represents the target pixel points in the wavelength；
3), with wavelength value as x-axis, it is with gray value after the pixel gray level Value Data of picture under having read all wavelengths then
Y-axis can describe the corresponding curve of spectrum of the pixel.
9. it is according to claim 8 for algae differentiate and algae proliferation non-destructive monitoring situation method, it is characterised in that
Described algae discrimination method is that the curve of spectrum for surveying pixel with target using the curve of spectrum of standard items compares；The target
Pixel point selection is located at the pixel within algae edge.
10. it is according to claim 1 differentiate for algae and algae proliferation non-destructive monitoring situation method, its feature exists
In the statistical method of the algae is after carrying out species discriminating to each frond, different colours to be used in spectrum micro-image
Different algal species are marked, the pixel being located within algae edge is calculated as a frond, and labeled as same color,
And count all kinds of same color frond numbers.
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