CN109124615B - Selective area high dynamic laser speckle blood flow imaging device and method - Google Patents
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Abstract
The invention discloses a selective area high dynamic laser speckle blood flow imaging device and a method, comprising a laser light source, a sample fixing table, a beam expander, a half-reflecting half-lens, a first CCD camera, a second CCD camera and a stepping motor, wherein the original speckle image is continuously acquired in a multi-exposure acquisition mode, and the defect that the original speckle image is acquired with fixed exposure time in the traditional laser speckle contrast imaging method is overcome. In addition, the invention provides a double imaging system, wherein the fixed focus system performs whole body blood flow imaging on a required imaging sample, the zoom system can perform optical amplification operation on a local area to obtain a blood flow contrast image with richer blood vessel information, and finally, the images obtained by the two optical systems are fused to obtain the blood flow contrast image with higher imaging resolution. The method has important significance in the fields of early diagnosis of diseases, disease analysis, dynamic monitoring of drug effects in vivo and the like.
Description
Technical Field
The invention relates to the field of optical imaging, in particular to a selective area high dynamic laser speckle blood flow imaging device and method.
Background
The laser speckle blood flow imaging technology is widely applied in the biomedical research field, and provides a full-field optical blood flow imaging method without scanning. Such imaging systems use laser light to illuminate a sample to generate a speckle signal, and then use a high sensitivity coupling element (CCD) to receive the speckle signal emitted from the object to obtain a full-field two-dimensional blood flow distribution image. The speckle signals can be divided into dynamic speckle and static speckle, the speckle signals with random jitter of the speckle signal intensity in the area where the scattering particles are located caused by the movement of the scattering particles are called dynamic speckle, whereas the speckle signals with no random jitter of the speckle signal intensity are called static speckle. Taking a contrast analysis method [ CN102429650A ] for laser speckle blood flow imaging as an example, the patent of the invention discloses an experimental device and a method for laser speckle imaging. The system irradiates laser beams on a sample to be measured, and continuously acquires speckle images reflected by N frames of measured objects at the same exposure time and frame interval time.
The exposure time is an important parameter of camera imaging, and the imaging of the sample by adopting the same exposure time obviously cannot meet the requirement of different parts on the required exposure time due to the difference of the thicknesses of different parts of the biological sample. Taking a laser speckle blood flow imaging method [ CN103330557A ] based on exposure time measurement as an example, the invention provides a novel imaging method aiming at the requirement of different parts of an imaged sample on required exposure time, and a series of exposure time is adopted to respectively and continuously collect the sample.
In the existing imaging device and method for acquiring original speckle images by utilizing multiple exposures, CCD cameras are used for continuously acquiring target monitoring points at different exposure times respectively, acquired image data are transmitted to a computer, the relation of the attenuation time of an autocorrelation function of scattering light intensity fluctuation is calculated by utilizing a formula and is fitted, finally, an exposure time value is obtained, and then the obtained exposure time is set for carrying out blood flow imaging on a detection object. The method uses a formula to calculate the optimal exposure time, and uses the exposure time to image the imaged sample. However, this method requires that the imaged sample should have a uniform thickness, and for some biological samples with different thicknesses, it is obviously impossible to meet the required exposure time requirements of different sites with only one optimal exposure time, and thus a high signal-to-noise ratio angiographic image cannot be obtained.
In addition, in the existing laser speckle blood flow imaging device, the original speckle image is mostly continuously acquired by adopting fixed exposure time or by obtaining the optimal exposure time under a series of exposure time gradients. However, in the whole-body blood flow imaging of an organism, since different parts of a biological sample often have different thicknesses and blood flow information of different parts is different, the requirement of different parts on the exposure time required for imaging may not be met by adopting a single exposure time.
Disclosure of Invention
Aiming at the defects of the existing laser speckle blood flow imaging technology, the invention provides a selective area high dynamic laser speckle blood flow imaging device and a selective area high dynamic laser speckle blood flow imaging method.
In order to achieve the above purpose, the invention adopts the following technical scheme.
The utility model provides a high dynamic laser speckle blood flow imaging device of selectable district, includes laser light source, sample fixed station, beam expander, half anti-half lens, first CCD camera, second CCD camera and step motor, wherein laser light source transmission laser beam passes through beam expander shines on the sample that awaits measuring on the sample fixed station, scattering particles in the sample that awaits measuring takes place the scattering and forms the speckle back to incident light and pass through half anti-half mirror beam split forms reflection facula and transmission facula, reflection facula is received by first CCD camera, transmission facula is received by second CCD camera, the second CCD camera is fixed on the step motor, step motor can be under the tiny removal of the control of driver, first CCD camera with second CCD camera passes through cable wire and peripheral processing terminal communication connection.
The first CCD camera and the second CCD camera both comprise CMOS sensor components.
The invention also provides a selective area high dynamic laser speckle blood flow imaging method, which comprises the following steps:
s1, fixing the whole device on a vibration isolation optical platform, opening a laser, and enabling a laser beam to act on a sample to be detected on a sample stage after being expanded by a beam expander;
s2, scattering particles in the sample to be detected scatter incident light, and scattered light generates speckle signals through random interference;
s3, one part of the speckle signal is reflected at the half-reflecting and half-transmitting mirror and is reflected to the fixed-focus CCD camera at the reflecting end, and the other part of the speckle signal is received by the zoom CCD camera at the transmitting end through the half-reflecting and half-transmitting lens;
s4, accurately focusing the imaged sample by controlling the stepping motor to slightly move the zoom CCD camera, so as to obtain a locally amplified blood flow radiography image;
s5, extracting characteristic values from blood flow information of the local amplified image and the original image, and performing image registration operation to determine the position of the amplified image in the original image;
s6, fusing the amplified local image with the whole body blood flow image through an image fusion algorithm, so as to obtain a whole body blood flow radiography image with high imaging resolution;
s7, respectively using two CCD cameras with exposure time t 1 ，t 2 ，……，t n Collecting m frames of original speckle images to obtain n x m frames of image sets with n exposure gradients in total, wherein P is 1 （m），P 2 （m），……，P n (m) represents the following. Wherein t is n For the maximum exposure time, then transmitting the acquired original speckle images under a series of exposure gradients to a processing terminal;
s8, respectively calculating different exposure times t i Laser speckle blood flow imaging parameters under the condition;
s9, respectively carrying out image fusion on the high-dynamic whole body blood flow images acquired by the first CCD camera by adopting an image fusion algorithm to acquire high-dynamic whole body blood flow contrast images, and carrying out fusion on the high-dynamic local blood flow images acquired by the second CCD camera to acquire high-dynamic local blood flow contrast images;
s10, positioning the amplified image in the region where the original image is located by adopting an image registration method according to the information of the large blood vessels in the amplified image of the selected region and the whole body image, and finally fusing the locally amplified image with the whole body image by utilizing an image fusion method to obtain a whole body blood flow radiography image with higher resolution.
The intensity of the collected speckle signals can be calculated by processing the information received by the terminal:
wherein,,
indicating speckle signal intensity, +.>
Scattered light intensity generated for scattering medium in background tissue, < >>
Is noise signal intensity, ++>
Is the dynamic speckle signal intensity in the imaged biological sample.
Performing fast Fourier transform on the formula (1) to obtain frequency domain distribution of speckle signals, setting a frequency domain window according to experimental conditions, performing inverse Fourier transform on a static signal at a low frequency and a dynamic signal at a high frequency respectively to obtain time domain static and dynamic signals, and calculating to obtain the concentration of dynamic scattering particles:
wherein,,
dynamic frequency domain signal corresponding to scattering particles representing motion, < >>
Representing static frequency domain signals corresponding to the background high scattering medium, wherein the acquisition frame rate is +.>
The filtering range is +.>
(x, y) is denoted as pixel position.
The invention adopts a multi-exposure acquisition mode to continuously acquire the original speckle image, and solves the defect that the original speckle image is acquired with fixed exposure time in the traditional laser speckle contrast imaging method. By acquiring a series of high dynamic range images in a dynamic range imaging system, then processing the images of each exposure gradient, extracting useful information of each image which is not lost in the normal exposure range through a post-processing algorithm, and reconstructing the useful information into an image with a high dynamic range.
In addition, the invention provides a double imaging system, wherein the fixed focus system performs whole body blood flow imaging on a required imaging sample, the zoom system can perform optical amplification operation on a local area to obtain a blood flow contrast image with richer blood vessel information, and finally, the images obtained by the two optical systems are fused to obtain the blood flow contrast image with higher imaging resolution.
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FIG. 1 is a schematic diagram of a high dynamic laser speckle blood flow imaging device of the present invention with an optional zone;
FIG. 2 is a flow chart of the data processing operation performed on an original speckle image in accordance with the present invention.
Detailed Description
For the purpose of facilitating a better understanding of the nature of the present invention by those of ordinary skill in the art, reference will now be made in detail to the following detailed description of the invention taken in conjunction with the accompanying drawings.
Referring to fig. 1, a high dynamic laser speckle blood flow imaging device for a selectable region comprises a laser light source 1, a sample fixing table 2, a beam expander 3, a half-reflecting mirror 4, a first CCD camera 5, a second CCD camera 6 and a stepping motor 7, wherein the whole device is firstly fixed on a vibration isolation optical platform, then a sample to be detected is fixed on the sample table 2, the laser light source 1 is opened to emit laser beams, the irradiation area of the laser is enlarged through the beam expander, the laser irradiation is more uniform, then the enlarged laser beams irradiate the sample to be detected, scattering particles in the sample to be detected scatter incident light, scattered light is randomly interfered in a far field to generate speckles, speckle signals are respectively received by imaging lenses at two ends after passing through the half-reflecting lenses, the collected fluorescent signals with specific wavelengths are focused on the CCD cameras, the CCD cameras convert the received optical signals into electric signals, the electric signals are stored in an acquisition card through a cable 8, and the acquired original speckle images are transmitted to a processing terminal 9 by the acquisition card for subsequent data processing operation.
The imaging system of the invention is a double imaging system. The side of the first CCD camera 5 is fixed, and is mainly used for imaging the whole body blood flow of an imaging sample as a fixed focus system of the device. The second CCD camera 6 is movable on the side as a zoom system of the present apparatus. Any region of the imaged sample may be selected for magnification. The second CCD camera 6 is fixed on the stepper motor 7, and in experiments, accurate focusing on the imaged sample can be achieved by controlling the micro motion of the stepper motor 7. By this operation of optically magnifying the local area, a more informative angiographic image of the blood vessel can be obtained. Because the local amplified image and the whole body range image have the same large blood vessels, the characteristic values are extracted by utilizing blood flow information existing in the local amplified image and the whole body range image, and the image registration operation is carried out, so that the position of the amplified image in the original image is found, and finally, the amplified local image and the whole body blood flow image are fused through an image fusion algorithm, so that the whole body blood flow radiography image with high imaging resolution is obtained.
The invention also provides a selective area high dynamic laser speckle blood flow imaging method, which comprises the following steps of:
s1, fixing the whole device on a vibration isolation optical platform, opening a laser, and enabling a laser beam to act on a sample to be detected on a sample stage after being expanded by a beam expander;
s2, scattering particles in the sample to be detected scatter incident light, and scattered light generates speckle signals through random interference;
s3, one part of the speckle signal is reflected at the half-reflecting and half-transmitting mirror and is reflected to the fixed-focus CCD camera at the reflecting end, and the other part of the speckle signal is received by the zoom CCD camera at the transmitting end through the half-reflecting and half-transmitting lens;
s4, accurately focusing the imaged sample by controlling the stepping motor to slightly move the zoom CCD camera, so as to obtain a locally amplified blood flow radiography image;
s5, extracting characteristic values from blood flow information of the local amplified image and the original image, and performing image registration operation to determine the position of the amplified image in the original image;
s6, fusing the amplified local image with the whole body blood flow image through an image fusion algorithm, so as to obtain a whole body blood flow radiography image with high imaging resolution;
s7, respectively using two CCD cameras with exposure time t 1 ，t 2 ，……，t n Collecting m frames of original speckle images to obtain n x m frames of image sets with n exposure gradients in total, wherein P is 1 （m），P 2 （m），……，P n (m) represents the following. Wherein t is n For the maximum exposure time, then transmitting the acquired original speckle images under a series of exposure gradients to a processing terminal;
s8, respectively calculating different exposure times t i Laser speckle blood flow imaging parameters under the condition;
s9, respectively carrying out image fusion on the high-dynamic whole body blood flow images acquired by the first CCD camera by adopting an image fusion algorithm to acquire high-dynamic whole body blood flow contrast images, and carrying out fusion on the high-dynamic local blood flow images acquired by the second CCD camera to acquire high-dynamic local blood flow contrast images;
s10, positioning the amplified image in the region where the original image is located by adopting an image registration method according to the information of the large blood vessels in the amplified image of the selected region and the whole body image, and finally fusing the locally amplified image with the whole body image by utilizing an image fusion method to obtain a whole body blood flow radiography image with higher resolution.
The original laser speckle image is acquired and the data processing operation process is as follows:
(1) Respectively calculating different exposure time t i Laser speckle blood flow imaging parameters under condition
At exposure time t i Under the condition of (1), the intensity of speckle signals acquired by each pixel point of the camera is assumed to be
Which contains the backgroundLight intensity, system noise, and dynamic speckle signal intensity carried by moving red blood cells. Therefore, the original speckle signal strength of any pixel received by the camera can be expressed as:
wherein,,
scattered light intensity generated for high scattering medium in background tissue,/->
Is noise signal intensity, ++>
Is the dynamic speckle signal intensity in the imaged biological sample.
To obtain a clearer blood flow contrast image, we also need to perform subsequent data processing operations on the original speckle image:
and performing fast Fourier transform on the formula to obtain the frequency domain distribution of the speckle signals. When the frequency domain distribution of the speckle signals is analyzed, the speckle signal intensity of the area where the scattering particles are located is randomly dithered due to the movement of the scattering particles, the corresponding speckle signals can be regarded as alternating current signals, and the frequency signals are distributed in the whole frequency domain mainly by superposition of the frequency signals. Under the condition of neglecting electronic noise, as no blood cells pass through the background tissue, the corresponding speckle signals can be regarded as direct current signals and are mainly distributed in the zero frequency region. Then setting a frequency domain window according to experimental conditions, and respectively carrying out inverse Fourier transform on the static signal at low frequency and the dynamic signal at high frequency so as to obtain time domain static and dynamic signals. Will be
Dynamic frequency domain signal corresponding to scattering particles representing motion, < >>
Representing static frequency domain signals corresponding to a background high scattering medium, wherein the acquisition frame rate is f, and the filtering range is +.>
. The imaging parameter MD is defined as the ratio of the dynamic signal intensity to the static signal intensity, i.e. the dynamic scattering particle concentration can be expressed as:
where (x, y) is denoted as pixel position,MD(x,y) The representation coordinates are%x,y) The pixel locations reconstruct the modulation depth values of the image. Laser speckle blood flow images under single exposure time conditions can thus be obtained separately.
(2) Reconstructing a multi-exposure image
Image fusion is carried out on the obtained blood flow contrast images under different exposure conditions by adopting an image fusion algorithm:
respectively the exposure time t 1 ，t 2 ，……，t n The m frames of images acquired under the condition are processed by the operation, and the processed images are represented by B (t 1 )，B(t 2 )，B(t 3 ), ……，B(t n ) Representing that a set of n images is obtained. We use here the image fusion method of the high frequency band. The high-frequency coefficient fusion adopts a principle of taking the absolute value of a pixel point, and a point with the large absolute value of the pixel point is selected as the pixel value of the fused image by comparing the absolute values of the pixel points of adjacent images. Finally, a blood flow radiography image with high dynamic range can be obtained.
(3) Optional region image magnification and registration
In the invention, a new method is introduced for amplifying the local area image to obtain a blood flow radiography image with more abundant blood vessel information. According to the information of large blood vessels in the selected region amplified image and the whole body image, the amplified image is positioned in the region where the original image is positioned by utilizing an image registration method, and finally the locally amplified image and the whole body image are fused by utilizing an image fusion method, so that a blood flow radiography image with higher imaging resolution is obtained.
The above embodiments are described in detail for the essence of the present invention, but the scope of the present invention is not limited thereto. It will be apparent to those skilled in the art that many improvements and modifications can be made without departing from the spirit of the invention, and it should be noted that these improvements and modifications fall within the scope of the appended claims.
Claims (4)
1. The method is characterized in that the method utilizes a selective area high-dynamic laser speckle blood flow imaging device, the selective area high-dynamic laser speckle blood flow imaging device comprises a laser light source, a sample fixing table, a beam expander, a half-reflection and half-lens, a first CCD camera, a second CCD camera and a stepping motor, wherein the laser light source emits laser beams to irradiate a sample to be detected on the sample fixing table through the beam expander, scattering particles in the sample to be detected scatter incident light and form speckles, and then the scattered particles are split by the half-reflection and half-transmission mirror to form reflection spots and transmission spots, the reflection spots are received by the first CCD camera, the transmission spots are received by the second CCD camera, the second CCD camera is fixed on the stepping motor, the stepping motor can move slightly under the control of a driver, and the first CCD camera and the second CCD camera are in communication connection with a peripheral processing terminal through a cable;
the selective area high dynamic laser speckle blood flow imaging method comprises the following steps:
s1, fixing the whole device on a vibration isolation optical platform, opening a laser, and enabling a laser beam to act on a sample to be detected on a sample stage after being expanded by a beam expander;
s2, scattering particles in the sample to be detected scatter incident light, and scattered light generates speckle signals through random interference;
s3, one part of the speckle signal is reflected at the half-reflecting and half-transmitting mirror and is reflected to the fixed-focus CCD camera at the reflecting end, and the other part of the speckle signal is received by the zoom CCD camera at the transmitting end through the half-reflecting and half-transmitting lens;
s4, accurately focusing the imaged sample by controlling the stepping motor to slightly move the zooming CCD camera, so as to obtain a locally amplified blood flow radiography image;
s5, acquiring m frames of original speckle images by two CCD cameras respectively with exposure time t1, t2, … … and tn to obtain n x m frames of image sets with n exposure gradients, wherein tn is the maximum exposure time, and then transmitting the acquired original speckle images under a series of exposure gradients to a processing terminal;
s6, calculating laser speckle blood flow imaging parameters under different exposure time ti conditions respectively;
s7, respectively carrying out image fusion on the high-dynamic whole body blood flow images acquired by the first CCD camera by adopting an image fusion algorithm to acquire high-dynamic whole body blood flow contrast images, and carrying out fusion on the high-dynamic local blood flow images acquired by the second CCD camera to acquire high-dynamic local blood flow contrast images;
s8, positioning the amplified local blood flow image in the region where the whole body blood flow image is located by adopting an image registration method according to the information of the large blood vessels in the amplified local blood flow image and the whole body blood flow image of the selected region, and finally fusing the amplified local blood flow image with the whole body blood flow image by utilizing an image fusion method to obtain a whole body blood flow radiography image with higher resolution.
2. A method of selective area high dynamic laser speckle blood flow imaging of claim 1, wherein: the first CCD camera and the second CCD camera both comprise CMOS sensor components.
3. A method of selective area high dynamic laser speckle blood flow imaging of claim 1, wherein:
the intensity of the collected speckle signals can be calculated by processing the information received by the terminal:
wherein,,
indicating speckle signal intensity, +.>
Scattered light intensity generated for scattering medium in background tissue, < >>
Is noise signal intensity, ++>
Is the dynamic speckle signal intensity in the imaged biological sample.
4. A method of selective area high dynamic laser speckle blood flow imaging as set forth in claim 3 wherein:
performing fast Fourier transform on the formula (1) to obtain frequency domain distribution of speckle signals, setting a frequency domain window according to experimental conditions, performing inverse Fourier transform on a static signal at a low frequency and a dynamic signal at a high frequency respectively to obtain time domain static and dynamic signals, and calculating to obtain the concentration of dynamic scattering particles:
wherein,,
dynamic frequency domain signal corresponding to scattering particles representing motion, < >>
Representing static frequency domain signals corresponding to the background high scattering medium, wherein the acquisition frame rate is +.>
The filtering range is +.>
(x, y) is denoted as pixel position.
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