CN110082350A - The microscopic imaging device and measurement method adaptively illuminated based on high-power LED lighteness - Google Patents

Abstract

A microscopic imaging device and measurement method based on high-power LED brightness adaptive illumination, the device includes: a computer, an LED control module, an LED array, a stage, a microscopic objective lens, a tube lens, and an optical detector. The LED units on the LED array sequentially illuminate the sample to be tested on the stage, and the illumination light passes through the sample to form diffracted light, which is collected by the microscope objective lens and lens barrel lens and then irradiates on the imaging plane of the optical detector, and Record this low-resolution intensity image. Using the iterative reconstruction algorithm can realize the rapid recovery of the high-resolution complex amplitude of the sample. By increasing the power of the LED and decomposing the LED light source in a multi-modal manner, the quality and accuracy of the microscopic imaging can be effectively improved. It is suitable for many fields in the fields of materials science and biomedicine. Optical microscopic imaging of such samples has broad application prospects.

Description

Microscopic imaging device and measurement method based on high-power LED brightness adaptive illumination

technical field

The invention relates to the field of quantitative phase measurement and optical microscopic imaging, in particular to a microscopic imaging device and measurement method based on high-power LED brightness adaptive illumination.

Background technique

The traditional optical microscopic imaging technology is mainly based on the principle of lens imaging. The local microscopic imaging of the sample to be tested is realized through the lens group. Due to the limitation of the numerical aperture of the objective lens, increasing the imaging resolution of the objective lens will reduce the size of the imaging field of view. Mutual constraints, difficult to balance. In order to solve this contradiction, the conventional microscope system mainly uses a precision motorized platform to scan a wide range of airspace, and uses software to stitch and fuse images in continuous field of view. However, this method relies on high-precision mechanical scanning components, which inevitably introduces mechanical errors while increasing the system cost, and affects the quality of microscopic imaging. In addition, the optical detector can only record the intensity information of the beam, but the phase information determined by the three-dimensional shape and refractive index of the sample is lost in the process of propagation and recording. Therefore, how to obtain accurate quantitative phase imaging has also become a challenge. An important proposition in the field of microscopy imaging. With the introduction of the ptychography iterative engine (PIE) phase recovery iterative algorithm and its application in the frequency domain, optical microscopes have been able to use LED array illumination and phase recovery algorithms to obtain quantitative phase imaging with high resolution and large field of view. However, there are still some problems in the traditional frequency-domain PIE microscopic imaging technology. The LED unit power used in the LED array suitable for microscopic imaging illumination is generally small, and the brightness is low. The high-frequency signal is too weak, resulting in the loss of dark field information, and the resolution of the image restored by the frequency domain PIE method is reduced; secondly, the brightness of the LED unit cannot be adjusted independently, which can neither correct the random deviation between the brightness of different LED units, but also easily cause the center The image collected under the area LED lighting is overexposed; in addition, the core problem of using LED array lighting is that the spatial coherence of the LED light source is poor, and the illumination light emitted is partly coherent light, and the frequency domain PIE phase recovery algorithm is based on the coherent imaging system. Therefore, imaging errors will be introduced in the process of phase recovery, which will reduce the resolution and imaging quality of the microscopic imaging system.

In summary, improving the lighting power of LED arrays, realizing adaptive adjustment of LED unit brightness, and correcting the reconstruction error introduced by the weak coherence of LED light sources through algorithms have become new challenges for quantitative phase measurement in the field of microscopic imaging. Advanced LED lighting device and phase recovery algorithm to promote the transformation of optical microscopy technology.

SUMMARY OF THE INVENTION

The purpose of the present invention is to provide a microscopic imaging device and measurement method based on high-power LED brightness self-adaptive lighting, through the use of high-power LED driven lighting and arbitrary adjustment of LED light source brightness distribution, the process of microscopic imaging light field propagation is enhanced The high-frequency signal in the medium improves the quality of the dark field image collected by the optical detector, and the LED illumination light is corrected by the algorithm through multi-modal decomposition, which eliminates the imaging error caused by the poor spatial coherence of the LED, and is suitable for materials Quantitative microscopic imaging of a variety of samples in science and biomedicine.

The technical solution adopted by the present invention to solve its technical problems is:

A microscopic imaging device based on high-power LED brightness adaptive lighting, comprising: a computer, an LED control module, an LED array, an object stage, a microscopic objective lens, a lens tube lens, and an optical detector, and the LED control module includes a single-chip microcomputer Development board, switching power supply circuit, LED driver chip, decoding relay, load resistor; the output end of the computer is connected with the input end of the LED control module through the serial communication interface; the single-chip microcomputer development board is connected through the analog signal I/O interface It is connected with the input end of the LED driver chip, and the single-chip microcomputer development board is connected with the input end of the decoding relay through the digital signal I/O interface; the current output end of the switching power supply circuit is connected with the decoding relay; the LED array The current input terminal is connected to the output terminal of the decoding relay through multiple wires, the current output terminal of the LED array is connected to the LED driver chip through a load resistor; the output terminal of the optical detector is connected to the input terminal of the computer; the The center of the LED array is on the optical axis of the microscope; the rear focal plane of the microscopic objective lens overlaps the front focal plane of the lens barrel lens, and the imaging plane of the optical detector overlaps the rear focal plane of the lens barrel lens. , place the sample to be tested on the stage, and adjust the position so that the sample is at the front focal plane of the microscope objective lens, forming an optical coherent imaging system.

The light emitted by the LED units in the i-th row and j-column of the LED array is irradiated on the sample to be tested on the stage, and the illumination light passes through the sample to be tested to form diffracted light, which is collected by the microscope objective lens, and then passed through the barrel lens The converging becomes parallel light again, and irradiates on the imaging plane of the optical detector, and the obtained light intensity distribution is recorded by the optical detector and saved to the computer.

A microscopic imaging measurement method based on high-power LED brightness adaptive illumination, the steps are as follows:

i. Configure the LED array lighting parameters and start the LED control module, the steps are as follows:

a) Define the shape of the LED array light source, select the shape of the LED light source as ring, circle or rectangle according to the imaging needs, and input the specific shape parameters into the graphical operation interface of the computer.

b) Calculate the parameters of the LED units to be lit according to the shape parameters of the input LED light source, including information such as the position, order, frequency, and brightness distribution of the LED units to be lit, and save them to the computer program memory, and then convert the information to a numerical value The form of the array is sequentially written into the serial communication interface of the computer according to the predetermined frequency and saved, waiting for the lower computer to read.

c) The single-chip development board continuously reads the array data from the serial communication interface, and splits and decodes the array data, and after identifying the LED unit coordinates, brightness and lighting frequency, writes it into the I/O of the single-chip development board port, waiting for the lower computer to read.

d) The decoding relay reads the column coordinate signal from the digital I/O port of the single-chip microcomputer development board; the LED driver chip reads the row coordinate signal and brightness value from the analog I/O port of the single-chip microcomputer development board.

e) The switching power supply circuit is turned on, the LED driving circuit is started, the current flows in from the input terminal of the switching power supply circuit, the current is stabilized through the switching power supply, and then the decoding relay is used for column gating, and the feedback flows through the LED array to the LED driving chip The terminal performs row gating, and adjusts the current value of the single channel according to the PWM signal to adjust the brightness of the LED unit.

ii. The original image of microscopic imaging is collected, and the LED array is used as the light source of the microscopic imaging system for illumination, and the LED units on the LED array are sequentially lit, and the LED units in the i-th row and j-column emit illumination light and irradiate the light on the stage On the sample to be tested, the illumination light passes through the sample and is collected and converged by the microscope objective lens and lens tube lens, and finally collected by the optical detector and recorded by the computer;

iii. Specific steps to achieve wavefront reproduction through computer iterative operations

a) First generate the high-resolution initial guess O ₀ (u, v) of the sample to be tested and the initial guess P(u, v) of the pupil function of the objective lens in the frequency domain, usually using the optical detector record when the illumination light is vertically incident The low-resolution image of is used as the high-resolution initial light intensity of the sample to be tested, and the phase is initialized to zero.

b) Multimodal decomposition of the LED unit, decomposing each LED unit into several sub-modes with different positions and irrelevant to each other, and using the decomposed sub-modes to replace the original LED lighting light for simultaneous illumination and light field spread.

c) Calculate the complex amplitude of the low-resolution image plane of each mode, corresponding to the incident angle of each sub-mode, use the pupil function of the microscope objective lens to intercept the spectral information in a certain sub-aperture in the initial high-resolution spectrum of the sample, and pass The inverse Fourier transform is propagated to the image plane to generate complex amplitudes of the low-resolution image plane corresponding to each mode.

d) Update the complex amplitude of the image plane corresponding to each mode, calculate the total light intensity value of the imaging plane when each mode is illuminated at the same time, based on the ratio of the image plane intensity value corresponding to each mode to the total light intensity value, use the optical detector The recorded low-resolution light intensity images update the complex amplitudes of the low-resolution image planes of each modality.

e) Update the high-resolution spectrum of the sample and the pupil aperture function of the microscope objective lens, calculate the low-resolution spectrum difference corresponding to each mode before and after the complex amplitude update of the image plane, and calculate the corresponding value of each mode based on the spectrum difference The spectral sub-aperture in the high-resolution spectrum of the sample is updated, and the pupil aperture function of the microscope objective is updated at the same time.

f) Repeat the calculation steps a to e until the update of the sub-aperture spectrum in the high-resolution spectrum of the object corresponding to all illumination angles is completed. Then iterate the above process until the high-resolution spectrum of the object converges, and perform an inverse Fourier transform on the reconstructed high-resolution spectrum to obtain the high-resolution complex amplitude of the object.

Technical effect of the present invention:

1) The device has a simple structure, low environmental requirements, and does not depend on a precise mechanical translation stage, which reduces the cost of the device and avoids mechanical errors; the phase recovery algorithm proposed in the invention can break through the limitation of the numerical aperture of the microscopic objective lens, and expand the field of view Field at the same time to improve the resolution of microscopic imaging, to achieve accurate quantitative phase measurement.

2) Compared with other optical microscopic imaging devices using phase recovery algorithm, the present invention greatly improves the illumination light power of the LED unit, and can arbitrarily adjust the brightness of a single LED unit according to the imaging needs, significantly increasing the LED unit’s large-angle illumination. The high-frequency information improves the accuracy of the image collected by the optical detector and the imaging quality of the frequency-domain PIE microscopic imaging method.

3) The present invention performs multimodal decomposition on a single LED unit, calculates and fits the distribution of LED lighting light closer to the real one, effectively reduces the influence of the weak coherence of the LED light source, significantly improves the imaging accuracy, and restores the obtained high resolution The complex amplitude image is closer to the real sample.

The present invention will be described in further detail below in conjunction with the accompanying drawings.

Description of drawings

Figure 1 is a schematic diagram of a microscopic imaging device based on high-power LED brightness adaptive illumination.

Fig. 2 is a schematic structural diagram of the LED control module assembly.

Fig. 3 is a schematic diagram of the LED array circuit structure.

Fig. 4 is a schematic flow chart of the microscopic imaging measurement method based on high-power LED brightness adaptive illumination according to the present invention.

Fig. 5 is a schematic flow chart of driving control of a high-power LED array.

Fig. 6 is a plane coordinate diagram of the LED unit.

Fig. 7 is a schematic diagram of multi-mode decomposition of the LED unit.

1. Computer, 2. LED control module, 3. LED array, 4. Stage, 5. Microscopic objective lens, 6. Lens tube lens, 7. Optical detector, 8. MCU development board, 9. Switching power supply circuit , 10. Switching inductance, 11. Field effect MOS tube, 12. Schottky diode, 13. Output capacitor, 14. LED driver chip, 15. Decoding relay, 16. Load resistor, 17. LED unit, 18. Plug pin connector.

Detailed ways

Referring to Fig. 1, the microscopic imaging device and measuring method based on high-power LED brightness adaptive lighting of the present invention are built on the basis of a microscope with an LED array as the lighting source, including a computer 1, which transmits lighting control instructions to the LED control Module 2, the LED control module 2 drives and controls the LED array 3 for illumination. Along the direction of the illumination light, there are LED array 3, stage 4, microscope objective lens 5, lens tube lens 6, and optical detector 7 in sequence. The center of the LED array 3 is on the optical axis of the microscope; the rear focal plane of the microscopic objective lens 5 overlaps with the front focal plane of the lens tube lens 6, and the imaging plane of the optical detector 7 overlaps with that of the lens tube lens 6. The rear focal planes are overlapped, and the sample to be tested is placed on the stage 4 during imaging, and the position is adjusted so that the sample is at the front focal plane of the microscopic objective lens 5, forming an optical coherent imaging system.

The output end of the computer 1 is connected with the input end of the LED control module 2 through a serial communication interface.

Referring to Fig. 2, described LED control module 2 comprises single-chip microcomputer development board 8, switching power supply circuit 9, LED driver chip 14, decoding relay 15, load resistance 16; The input terminals of the board 8 are connected, and the communication baud rate of the serial communication interface is set to 9600. The microcontroller development board 8 described in the present embodiment adopts the Arduino uno R3 development board based on ATmega328, and the development board contains 14 I/O ports, including 8 digital ports and 6 PWM analog ports.

The single-chip microcomputer development board 8 is connected with the computer 1 through a USB interface and receives power supply, and the input voltage value is 5V. The storage space of the Arduino development board is composed of the internal storage space integrated by its main control chip, including Flash, SRAM and EEPROM. The Flash capacity is 32KB, of which 0.5KB is used as the BOOT area for storing the boot program, realizing the function of downloading the main control program through the serial port; the other 31.5KB is used for storing the serial port information identification program and the LED driver control program. The capacity of the SRAM is 2KB, and as the MCU memory, the storage space therein is temporarily occupied when the MCU performs calculation work. The EEPROM has a capacity of 1KB and is a programmable read-only memory, and the data therein will not be lost when the Arduino is powered off or reset.

The switching power supply circuit 9 includes a switching inductor 10 , a field effect MOS transistor 11 , a Schottky diode 12 and an output capacitor 13 . The output current I _ouT of the switching power supply circuit 9 is equal to the sum of the operating currents of each channel, that is, I _OUT =∑ _i I _i , where I _i is the operating current of each channel from IFB ₁ to IFB _M. The output voltage V _OUT of the switching power supply circuit 9 is determined by the input voltage V _IN and the conduction ratio D of the field effect MOS transistor 11, and the calculation formula is

The operating current I _L of the switching inductor 10 is determined by the input voltage V _IN , the output voltage V _OUT and the output current I _OUT , and its calculation formula is Where η is the energy conversion efficiency of the circuit.

The operating current I _Q of the field effect MOS transistor 11 is determined by the output current I _OUT and the switch conduction ratio D, and its calculation formula is The working voltage V _Q of the field effect MOS transistor 11 is equal to the output voltage V _OUT .

The capacitance C _OUT of the output capacitor 13 is between 22 μF and 220 μF, specifically determined by the output current I _OUT , switch conduction ratio D, ripple voltage V _RIPPLE and boost switching frequency f _SW , and its calculation formula is:

In this embodiment, the LED driver chip 14 adopts the TPS61196-Q1 chip, and the various circuit parameters selected by this chip in this embodiment are shown in the following table:

The LED driver chip 14 is driven based on the switching power supply circuit 9, and includes six current sink paths, all of which can perform independent PWM current regulation, and each path can be adjusted within 50mA to 500mA, while the PWM signal input terminal can be adjusted at 0 The driving control can be carried out within the range of 5V, and the total load of the circuit can bear up to 300W, which is enough to support the lighting power required by the microscopic imaging device and measurement method based on high-power LED brightness adaptive lighting of the present invention. The six-way PWM signal input end of the LED driver chip 14 is connected with the six-way PWM analog I/O port of the single-chip microcomputer development board 8, and the signal enable end EN of the LED driver chip 14 is connected with the Vin end of the single-chip microcomputer development board 8, and the LED driver chip The ground terminal GND of 14 is connected to the GND of the single-chip microcomputer development board 8; the current feedback terminal of the LED driver chip 14 is connected to the gate g of the field effect MOS transistor 11 of the switching power supply circuit 9 .

The decoding relay 15 includes a decoding circuit and a relay circuit. In this embodiment, the decoding circuit adopts a 3-8-wire decoder, and the relay circuit adopts optocoupler isolation. The specific parameters of the decoding relay 15 are shown in the following table Show:

The three signal input terminals of the decoding relay 15 are connected with the digital I/O ports of the single-chip microcomputer development board 8 , and the current input terminals of the decoding relay 15 are connected with the cathode of the Schottky diode 12 in the switching power supply circuit 9 .

The resistance value R _i of the load resistor 16 is determined by the output voltage V _OUT of the switching power supply circuit 9, the LED operating voltage V _{LED(i, j)} and the LED operating current I _i , and its calculation formula is Among them, i and j are the row and column coordinate numbers of the LED unit respectively. In this embodiment, all load resistors 16 have a resistance value of 150Ω and a rated power of 25W.

Referring to FIG. 3 , the LED array 3 includes several LED units 17 and two pin connectors 18 , and all the LED units 17 are equidistantly arranged to form an M×N LED matrix. In this embodiment, the LED array 3 includes a total of 36 LED units 17 in 6 rows and 6 columns, and the distance between rows and columns of every two adjacent LED units 17 is 11.14 mm; all LED units 17 are red LEDs, and the The typical wavelength is 625nm, and the rated power is 1W~3W; all LED units 17 are arranged in a mesh circuit, the jth column is selected as a channel, and the ith row is selected as a high level, and the LEDs in the ith row and jth column can be lit. LED unit 17, the brightness of the LED unit 17 depends on the current value of the i-th row path. The pin connector 18 is a 6-row plug-in connector with a pitch of 2.54mm. The specific parameters of the LED array 3 adopted in the present embodiment are shown in the following table:

The six current input ends of the LED array 3 are connected to the current output ends of the decoding relay 15 , and the six current output ends of the LED array 3 are connected to the feedback input end of the LED driver chip 14 through the load resistor 16 .

After the LED unit 17 in the i-th row and the j-th column in the LED array 3 is turned on, it emits red illumination light and irradiates the sample to be tested on the stage 4, and the diffracted light passing through the sample to be tested is microscopically The objective lens 5 and the barrel lens 6 collect and converge, and finally irradiate on the imaging plane of the optical detector 7 to form the original microscopic image of the sample to be tested, which is saved by the computer 1 and processed by the next phase recovery algorithm.

In this embodiment, the microscopic objective lens 5 adopts a microscopic objective lens with 10 times magnification and a numerical aperture of 0.25. The output end of described optical detector 7 is connected with the input end of computer 1 through standard C port; Described optical detector 7 adopts monochrome CCD camera, and its resolution is 1600 x 1200, and pixel size is 5.5 μ m, frame rate The black-and-white image of the sample to be tested under monochromatic LED lighting can be recorded and transmitted to the computer 1 for real-time imaging.

Referring to Fig. 4, the present invention is based on the microscopic imaging measurement method of high-power LED brightness adaptive illumination, and its steps are as follows:

i. Referring to Figure 5, configure the LED array lighting parameters and start the LED control module 2, the steps are as follows:

a) Define the shape of the LED array light source, select the shape of the LED light source as rectangle, circle or ring according to the imaging needs, and input the specific shape parameters into the graphical operation interface of the computer. First, establish a coordinate system for parameter calculation in the computer, see Figure 6, take the lower left corner of the LED array as the coordinate origin, the lower boundary is the x-axis, the right direction is the positive direction of the x-axis, the left boundary is the y-axis, and the upward direction is y In the positive direction of the axis, a plane Cartesian coordinate system is established. The unit length of the rectangular coordinate system is 1mm, the full scale values of the x-axis and the y-axis are both length, and length is the side length value of the LED array. Assume that the center coordinates of the first LED unit are (x ₀ , y ₀ ), and the distance between two adjacent LED units is d.

When the shape of the LED light source is selected as a rectangle, it is necessary to input the length m, width n and center coordinates (a, b) of the rectangle into the computer. The range of input values m and n is: 0<m, n<length, where length is the side length value of the LED array; the range of input values a and b is: m/2<a<length-m/ 2, n/2<b<length-n/2.

When the shape of the LED light source is selected as a circle, you need to input the radius r of the circle and the coordinates (a, b) of the center of the circle in the computer. The value range of the input value r is: 0<r<length/2, where length is the LED The side length value of the array; the range of input values a and b is: r<a, b<length-r. When the shape of the LED light source is selected as a ring, it is necessary to input the ring inner diameter r1, the ring outer diameter r2 and the coordinates (a, b) of the center of the circle in the computer. The value range of the input values r1 and r2 is: 0<r1<r2≤length/ 2; The range of input values a and b is: r2<a, b<length-r2.

b) Calculate the parameters of the LED unit to be lit according to the input shape parameters of the LED light source. First, calculate the position number of the LED unit to be lit. Assume that the position coordinates of the LED unit to be lit are (x, y), and the position number is (i , j), where i is the row number and j is the column number.

When the shape of the LED light source is selected as a rectangle, the coordinate range of the LED unit to be lit is: am/2≤x≤a+m/2, bn/2≤y≤b+n/2, where x and y are LED Unit abscissa and ordinate. Round the obtained coordinate range, and calculate the row and column numbers of the LED units to be lit according to the rounded coordinate range: In order to round down the symbol and to ensure the approximation of the boundary of the rectangular domain, the row and column numbers of the LED units are rounded up in the formula.

When the shape of the LED light source is selected as a circle, the coordinate range of the LED unit to be lit is: (xa) ² +(yb) ² ≤ r ² , where x and y are the abscissa and ordinate of the LED unit, respectively. Round the obtained coordinate range, and calculate the row and column numbers of the LED units to be lit according to the rounded coordinate range: in i' and j' are the row number and column number of the LED unit in the center of the circle, respectively. In order to ensure the approximation of the circle boundary, the row and column numbers of the LED unit are rounded up.

When the shape of the LED light source is selected as a ring, the coordinate range of the LED unit to be lit is: r1 ² ≤(xa) ² +(yb) ² ≤r2 ² , where x and y are the abscissa and ordinate of the LED unit, respectively. Round the obtained coordinate range, and calculate the row and column numbers of the LED units to be lit according to the rounded coordinate range: in i' and j' are the row number and column number of the LED unit in the center of the ring area, respectively. In order to ensure the approximation of the ring area boundary, the row and column numbers of the LED unit are rounded up in the formula.

Secondly, calculate the brightness value of the LED unit that needs to be lit. In order to ensure that the high-frequency information is not lost, the brightness of the LED unit is set to a linear growth mode from low to high from the center of the shape of the LED light source to the boundary. In this embodiment, according to the 8-bit output of the I/O digital port of the Arduino development board used, assuming that the brightness value of the LED unit is lightness, the brightness of the central LED unit is set to 127, and the brightness of the edge LED unit is 255; The distance of the central LED unit is the standard, and the value is linearly taken in the integer range of [127, 255].

When the shape of the LED light source is selected as a rectangle, the brightness value of the LED unit whose position coordinates are (x, y) is: Where max{...} is the maximum value in parentheses, is the rounding down symbol.

When the shape of the LED light source is selected as a circle, the brightness value of the LED unit whose position coordinates are (x, y) is:

When the shape of the LED light source is selected as a ring, the brightness value of the LED unit whose position coordinates are (x, y) is:

Finally, input the lighting frequency f of the LED unit in Hz, that is, the lighting time interval between every two LED units is delay, The unit is ms.

After the calculation of the LED unit parameter information is completed, a four-digit one-dimensional array comdata[4] is generated, and its data format is an integer int, and the array is assigned a value, so that

comdata[0]=lightness, indicating the brightness value of the LED unit

comdata[1]=delay, indicating the lighting interval of the LED unit

comdata[2]=i, indicates the row number of the LED unit

comdata[3]=j, indicates the column number of the LED unit

After the assignment is completed, write the numerical array comdata[4] into the serial communication interface of computer 1 in sequence according to the established serial communication frequency and save it, waiting for the lower computer to read.

c) The single-chip microcomputer development board 8 continuously reads the array data from the serial communication interface. In this embodiment, the communication baud rate of the serial communication interface is first set to 9600, and the frequency of updating the array data by the serial communication interface is set to be f. Generate a row port array RowPin[M] and a column port array ColPin[N] in the single-chip microcomputer, where M is the number of rows of LED units 17 in the LED array 3, and N is the number of ports at the signal input end of the decoding relay 15. Then assign values to the row port array and the column port array to establish a one-to-one mapping between the row and column signals and the I/O ports of the microcontroller development board, so that RowPin[M]={...}, RowPin[0] to RowPin[ M] is the program number of the 8 analog output ports of the single-chip development board in turn; Let ColPin[N]={...}, ColPin[0] to ColPin[N] are the program numbers of the 8 digital output ports of the single-chip development board in turn.

When the MCU receives the serial port communication status from computer 1 as the true value, it reads the array comdata[4] transmitted by the serial port, splits and reads the array, and identifies the brightness, lighting frequency f and Row and column number (i, j). At the same time, the row signal array row[M] and the column signal array column[N] are generated in the single-chip microcomputer, and the two signal arrays are assigned and input during the lighting period of one LED unit.

Row signal assignment: assign the analog output port whose program number is RowPin[i] as lightness, that is, let row[RowPin[i]]=lightness, and assign 0 to other elements of the row signal array row[M].

Column signal assignment: convert the decimal column number j into an N-bit binary number through decimal conversion, and assign it to the column array column[N], that is, column[N]=j ₍₂₎ , where j ₍₂₎ is the decimal number j The N-bit binary representation of . Then assign the digital output port whose program number is ColPin[N] to column[N] in sequence.

Set the assignment waiting time to delay, so that During the waiting time, the lower computer performs the data reading operation by connecting to the I/O port of the single-chip microcomputer. After the waiting time passes, the assignment operation at this stage ends and the next LED unit lighting period begins.

d) During the lighting cycle of an LED unit, the decoding relay 15 reads the column coordinate signal from the digital I/O port of the single-chip microcomputer development board 8, and the single-chip microcomputer development board 8 transmits the N-bit column signal array column through the digital output port The data in [N] is transmitted to the input terminal of the decoding relay 15, and then the decoding relay 15 decodes the binary array column[N] into decimal data j through base conversion, and j is the column gate signal of the LED array.

Simultaneously, the LED driver chip 14 reads the row coordinate signal and brightness value from the analog I/O port of the single-chip microcomputer development board 8, and the said single-chip microcomputer development board 8 transmits the data in the M-bit line signal array row[M] through the analog output port To the PWM input terminal of the LED driver chip 14, the array data includes the row strobe signal and the brightness value of the LED unit at the same time.

e) Connect the switching power supply circuit 9 to the DC regulated power supply, the LED drive circuit starts, and the LED control module 2 officially starts working. The output voltage value of the DC regulated power supply is 30V, and the maximum current value is 5A. The operation of the switching power supply circuit 9 is divided into a charging process and a discharging process.

Charging process: When a forward voltage of an appropriate threshold is applied between the gate and the source of the field effect MOS transistor 11, the drain d of the field effect MOS transistor 11 and the source s are turned on, and the current flows out of the DC stabilized power supply and then flows through the switching inductor 10. The field effect MOS transistor 11 reaches the negative pole of the DC stabilized voltage power supply. During this process, the current on the inductor increases linearly and stores energy, while the Schottky diode 12 is in the reverse-bias cut-off state at this time, and the output capacitor 13 discharges to the output terminal by relying on the electric energy stored in the previous discharge process to maintain the load operation.

Discharging process: when the forward voltage between the gate and the source of the field effect MOS transistor 11 is canceled, the drain d of the field effect MOS transistor 11 and the source s are turned off, and the current flows through the switch inductor 10 after flowing out from the DC stabilized voltage supply. The Schottky diode 12 and the output capacitor 13 reach the negative pole of the DC stabilized power supply. During this process, the DC stabilized power supply and the switching inductor 10 jointly charge the output capacitor 13 and discharge to the output terminal to maintain the load operation.

The charging and discharging processes are carried out alternately and reach a steady state, that is, in a complete cycle, the electric energy stored in the switching inductor 10 during the charging process should be equal to the electric energy released during the discharging process.

The output voltage of the switching power supply circuit 9 is determined by the input voltage and the conduction ratio of the field effect MOS transistor 11:

In the formula, V _OUT is the boosted output voltage, V _IN is the boosted input voltage, T is the full working cycle of the switch, T _ON is the conduction time of the switch, and D is the conduction ratio of the switch.

The switching power supply circuit 9 supplies power to the LED array 3 through the decoding relay 15 and the LED driver chip 14, and realizes LED control lighting through column gating and row gating.

Column strobe: After the decoding relay calculates the column strobe signal j, the jth column circuit selected by the relay is turned on, and the other channels are turned off. Only the jth column LED unit has current input, that is, the column strobe is realized .

Row strobe: After the PWM input terminal of the LED driver chip 14 receives the row strobe signal from the microcontroller development board 8, it adjusts the current of each channel at the load feedback end according to the PWM input parameters of each channel, and the unselected channel circuits are blocked. turn off, the current value of the i-th row being gated is adjusted to Where V _LED is the conduction voltage drop of the LED unit, V _Out is the output voltage of the switching power supply, and R _i is the load resistance of row i.

After the switching power supply circuit 9 outputs a steady-state current, it supplies power to the LED array 3 through the column gating function of the decoding relay 15, and flows through the LED unit 17 on the LED array 3, through the load feedback of the load resistor 16 and the LED driver chip 14. Connected to each other, using the PWM control of the LED driver chip 14 to realize row gating, forming a complete closed LED driver control circuit, which can realize single-point control and self-adjustment of the brightness of the high-power LED unit, and meet the requirements of the present invention based on the self-adaptive brightness of the high-power LED. Illuminated microscopic imaging devices and measurement methods are needed.

ii. The original image of microscopic imaging is collected, and the successfully driven LED array 3 is used as the light source of the microscopic imaging system for illumination, and the LED units 17 on the LED array 3 are sequentially lit according to a predetermined order, and the illumination light emitted by the LED unit 17 is irradiated to the waiting room. After the sample is measured, it is enlarged and imaged by the microscope objective lens 5 and the lens tube lens 6, and finally collected by the optical detector 7, and the computer 1 records and saves the low-resolution original images of the sample under all LED lighting

iii. Specific steps to achieve wavefront reproduction through computer iterative operations

a) First generate the high-resolution initial guess O ₀ (u, v) of the sample to be tested and the initial guess P(u, v) of the pupil function of the objective lens in the frequency domain, usually using an optical detector 7 when the illumination light is vertically incident The linear interpolation of the recorded low-resolution images is used as the high-resolution initial light intensity of the sample to be tested, and the phase is initialized to zero. The initial guess of the microscope objective pupil function P(u, v) is:

Where u and v are the frequency domain coordinates of the light field, and f ₀ is the cut-off frequency of the microscopic objective lens 5 .

b) Multimodal decomposition of the LED unit, decomposing each LED unit into several sub-modes with different positions and irrelevant to each other, each sub-mode has a different illumination angle relative to the sample, so the respective corresponding sample spectral aperture centers The coordinates are also different. Assuming that the plane coordinates of the LED unit in row i, column j are (x, y), and the distance between its sth sub-mode and the center of the LED unit is (Δx _s , Δy _s ), the sample spectrum corresponding to this sub-mode The coordinates of the center of the aperture are Calculated as follows:

Where h represents the vertical distance between the LED array and the sample, and λ represents the wavelength of the LED illumination light. The decomposed sub-modes are used to replace the original LED illumination light to illuminate the sample and propagate the light field at the same time, and the transmitted light field behind the sample is calculated according to the superposition of several sub-modes. c) Calculating the complex amplitude of the low-resolution image plane of each mode, corresponding to the incident angle of each sub-mode, using the pupil function of the microscopic objective lens 5 to intercept the spectral information in a certain sub-aperture in the initial high-resolution spectrum of the sample, the first The low-resolution spectrum of the intercepted sample corresponding to the s sub-modes is Its calculation formula is:

The superscript k indicates the kth iteration, (i, j) indicates the row and column coordinates of the LED unit, and s indicates the sth sub-mode. Indicates the high-resolution spectral subaperture of the sample. Then perform an inverse Fourier transform on the low-resolution spectrum to obtain the complex amplitude of the low-resolution image plane corresponding to each sub-mode which is Represents the inverse Fourier transform.

d) Update the complex amplitude of the image plane corresponding to each mode, and calculate the total light intensity of the imaging plane when each mode is illuminated at the same time The calculation formula is:

In the formula, |...| means to find the modulus of a two-dimensional complex matrix;

Then, based on the ratio of the image plane intensity value corresponding to each mode to the total value of light intensity, the low-resolution light intensity image recorded by the optical detector 7 is used to update the low-resolution image plane complex amplitude of each mode, and update the formula as follows:

in is the complex amplitude of the low-resolution image plane updated for each mode, is the light intensity image recorded by the optical detector 7 when the i-th row and the j-th column are illuminated by the LED.

e) Update the high-resolution spectrum of the sample and the pupil function of the microscope objective lens 5, first calculate the low-resolution spectrum difference corresponding to each mode before and after the complex amplitude update of the image plane Its calculation formula is as follows:

In the formula Expressed as a Fourier transform.

Then update the spectral sub-aperture in the high-resolution spectrum corresponding to each mode based on the spectral difference, and the update formula is:

In the formula Indicates the updated high-resolution spectral sub-aperture of the sample; the superscript * indicates the conjugate function of the function; |...| _max indicates the maximum value of the modulus of the two-dimensional complex matrix; at the same time, the pupil function of the objective lens is updated , the update formula is:

P′ ^k (u, v) represents the updated pupil function of the microscope objective lens.

f) Repeat the calculation steps a to e until the update of the sub-aperture spectrum in the high-resolution spectrum of the object corresponding to all illumination angles is completed. Then iteratively repeat the above process until the high-resolution spectrum O′ _k (u, v) of the object converges, and perform an inverse Fourier transform on the reconstructed high-resolution spectrum, and finally obtain the high-resolution complex amplitude o′ _k of the object (x, y), i.e.

Claims (17)

1. a kind of microscopic imaging device adaptively illuminated based on high-power LED lighteness, comprising: computer (1), LED control mould Block (2), LED array (3), objective table (4), microcobjective (5), tube lens (6), optical detector (7)；

The input terminal of the computer (1) is connected with the output end of optical detector (7), and the output end of the computer (1) is logical It crosses serial communication interface to be connected with the input terminal of LED control module (2), the output end and the LED of LED control module (2) The input terminal of array (3) is connected, and the center of the LED array (3) is on microscopical optical axis；The microcobjective (5) Back focal plane and the front focal plane of tube lens (6) overlap, imaging plane and tube lens (6) of the optical detector (7) Back focal plane overlaps, and sample to be tested is placed on objective table (4) by when imaging, and adjusting position makes sample be in microcobjective (5) front focal plane constitutes optical coherence imaging systems.

2. the microscopic imaging device according to claim 1 adaptively illuminated based on high-power LED lighteness, feature are existed In the LED control module (2) includes microcontroller development board (8), switching power circuit (9), LED drive chip (14), decoding Relay (15) and load resistance (16)；

The input terminal of the microcontroller development board (8) is connected with the output end of computer (1) by USB interface and receives confession Electricity；The microcontroller development board (8) is connected by analog signal I/O interface with the signal input part of LED drive chip (14)；Institute The end Vin for stating microcontroller development board (8) is connected with the signal enable end EN of LED drive chip (14)；The microcontroller development board (8) GND ground terminal is connected with the GND ground terminal of LED drive chip (14)；The microcontroller development board (8) passes through digital signal I/O Interface is connected with the signal input part of decoding relay (15)；The switch control terminal and LED of the switching power circuit (9) drive Chip (14) is connected；The current output terminal of the switching power circuit (9) is connected with the current input terminal of decoding relay (15)； The current input terminal of the LED array (3) is connected by multichannel conducting wire with the output end of decoding relay (15)；It is LED gusts described The current output terminal of column (3) is connected by load resistance (16) with LED drive chip (14).

3. the microscopic imaging device according to claim 1 or 2 adaptively illuminated based on high-power LED lighteness, feature It is, the LED array (3) includes several LED units (17) and two contact pin connectors (18), all adjacent LED units (17) spacing is equal, and constitutes the LED matrix of M x N；LED unit (17) quilt that the i-th row jth arranges in the LED array (3) After lighting, illumination is issued on the sample to be tested being located on objective table (4), it is micro- through the diffraction light of sample to be tested Object lens (5) and tube lens (6) are collected and converge, and are finally radiated on the imaging plane of optical detector (7), formed to The original micro-imaging of sample is saved by computer (1) and is rebuild by iterative algorithm the high-resolution complex amplitude of sample.

4. the microscopic imaging device according to claim 1 or 2 adaptively illuminated based on high-power LED lighteness, feature It is, the switching power circuit (9) includes switched inductors (10), field-effect metal-oxide-semiconductor (11), Schottky diode (12), defeated Capacitor (13) out；

The output end of the switched inductors (10) is connected with the drain electrode d of field-effect metal-oxide-semiconductor (11)；The switched inductors (10) it is defeated Outlet is connected with the anode of Schottky diode (12)；The source electrode s of the field-effect metal-oxide-semiconductor (11) is connected with GND ground terminal；It is described The grid g of field-effect metal-oxide-semiconductor (11) is connected with the LED drive chip (14)；The cathode of the Schottky diode (12) with The input terminal of output capacitance (13) is connected；The cathode of the Schottky diode (12) and the electric current of the decoding relay (15) Input terminal is connected；The output end of the output capacitance (13) is connected with GND ground terminal.

5. the microscopic imaging device according to claim 1 or 2 adaptively illuminated based on high-power LED lighteness, feature It is, the resistance value R of the load resistance (16)_iBy the output voltage V of switching power circuit (9)_OUT, LED operation voltage V_{LED (i, j)}And LED operation electric current I_iIt is determined, its calculation formula is Wherein i and j is respectively LED unit (17) column locations number.

6. a kind of utilize any microscopic imaging device adaptively illuminated based on high-power LED lighteness of claim 1-5 The micro-imaging measurement method of progress, which comprises the following steps:

Step 1, LED array (3) lighting parameter is configured, and starts LED control module (2)；

Step 2, micro-imaging original image acquires: it is illuminated using the light source of LED array (3) as micro imaging system, according to The secondary LED unit (17) lighted on LED array (3), the LED unit (17) of the i-th row jth column issue illumination and are being located at load On sample to be tested on object platform (4), illumination light is collected and converges through microcobjective (5) and tube lens (6) after passing through sample It is poly-, it is finally acquired by optical detector (7), and record preservation is carried out by computer (1)；

Step 3, the reconstruction for realizing sample high-resolution complex amplitude is calculated by being iterated in computer (1).

7. the microscopic imaging device according to claim 6 adaptively illuminated based on high-power LED lighteness carries out micro- Imaging measurement method, it is characterised in that in step 1, specifically includes the following steps:

1.1) it defines LED array (3) light source form: being needed to select LED light source shape for cyclic annular, round shape or rectangle according to imaging, And light source form parameter is inputted to the graphical operation interface of computer (1)；

1.2) LED unit (17) parameter for needing to light is calculated according to the LED light source form parameter of input, including need to light The information such as position, sequence, frequency and the Luminance Distribution of LED unit (17), and save to computer (1) program internal memory, then will The information is sequentially written in computer (1) serial communication interface according to set frequency in the form of numerical value array and saves, under waiting Position is machine-readable to be taken；

1.3) microcontroller development board (8) persistently reads array data from serial communication interface, and carries out fractionation solution to array data After identification obtains LED unit (17) coordinate, brightness and illumination frequency information, and the end I/O of microcontroller development board (8) is written in code Mouthful, wait slave computer to read；

1.4) digital I/O mouthful reading column coordinate signals of the decoding relay (15) from microcontroller development board (8)；LED drive chip (14) from I/O mouthfuls of reading row coordinate signals of the simulation of microcontroller development board (8) and brightness value；

1.5) switching power circuit (9) is connected, LED drive circuit starting, input terminal stream of the electric current from switching power circuit (9) Enter, carry out current stabilization by Switching Power Supply, carry out column selection using decoding relay (15) and lead to, flows through LED array (3) and reach LED The feedback end of driving chip (14) carries out capable gating, and adjusts single pass current value according to pwm signal to adjust LED unit (17) brightness.

8. the microscopic imaging device according to claim 6 adaptively illuminated based on high-power LED lighteness carries out micro- Imaging measurement method, it is characterised in that in step 3, specifically include following calculating process:

3.1) the high-resolution initial guess O of sample to be tested is generated in a frequency domain₀At the beginning of (u, v) and microcobjective (5) pupil function Begin to guess P (u, v), the low-resolution image recorded using optical detector (7) when illumination light vertical incidence is as sample to be tested High-resolution initial beam intensity, be zero by initialisation phase；

3.2) multi-modal decomposition is carried out to LED unit (17), it is different and mutual that each LED unit (17) is decomposed into several positions Incoherent submodule state, and substitute original LED illumination light with several submodule states after decomposition while carrying out illumination and light field It propagates；

3.3) the low resolution image planes complex amplitude of each mode is calculated, the incidence angle of corresponding each submodule state utilizes microcobjective (5) Pupil function interception sample initial high resolution frequency spectrum in spectrum information in certain sub-aperture, and pass through inverse Fourier transform biography Image planes are multicast to, low resolution image planes complex amplitude corresponding to each mode is generated；

3.4) the corresponding image planes complex amplitude of each mode, the light intensity total value of imaging plane, base when calculating each mode while illuminating are updated In the ratio of each mode corresponding image planes intensity value and light intensity total value, the low resolution light being recorded using optical detector (7) Strong image is updated the low resolution image planes complex amplitude of each mode；

3.5) the pupillary aperture function of the high resolution spectrum of more new sample and microcobjective (5) calculates image planes complex amplitude and updates Low resolution spectrum difference corresponding to each mode in front and back, and based on spectrum difference to each mode corresponding sample high-resolution frequency Frequency spectrum sub-aperture in spectrum is updated, while being updated to the pupillary aperture function of microcobjective (5)；

3.6) step 3.1) is computed repeatedly to step 3.5), until completing object high-resolution frequency corresponding to all light angles Compose the update of sub-aperture frequency spectrum.Then iteration above procedure, until the high resolution spectrum of object is restrained, to rebuilding To high resolution spectrum make inverse Fourier transform, obtain the high-resolution complex amplitude of object.

9. the microscopic imaging device according to claim 7 adaptively illuminated based on high-power LED lighteness carries out micro- Imaging measurement method, it is characterised in that in step 1.1):

When selecting LED light source shape for rectangle, length m, width n and the center of the input rectangle in computer (1) are needed Coordinate (a, b).The value range of input value m and n are as follows: 0 < m, n < length, wherein length is the side length of LED array (3) Value；The value range of input value a and b are as follows: m/2 < a < length-m/2, n/2 < b < length-n/2；

When it is round for selecting LED light source shape, need in computer (1) the circular radius r of input and central coordinate of circle (a, B), the value range of input value r are as follows: 0 < r < length/2, the value range of input value a and b are as follows: r < a, b < length- r；

When it is annular for selecting LED light source shape, input circular inside diameters r1, annular outer diameter r2 and circle in computer (1) are needed Heart coordinate (a, b), the value range of input value r1 and r2 are as follows: 0 < r1 < r2≤length/2；The value range of input value a and b Are as follows: r2 < a, b < length-r2.

10. the microscopic imaging device according to claim 7 adaptively illuminated based on high-power LED lighteness carries out micro- Imaging measurement method, it is characterised in that in step 1.2),

LED unit (17) Position Number for needing to light is calculated, detailed process is as follows:

Assuming that the side length of LED array (3) is length, the coordinate of first LED unit (17) is (x₀, y₀), two adjacent LEDs Spacing between unit (17) is d；LED unit (17) position coordinates for needing to light are (x, y), and Position Number is (i, j), Middle i is row number, and j is column number；

When selecting LED light source shape for rectangle, LED unit (17) coordinate range for needing to light are as follows: a-m/2≤x≤a+m/2, b-n/ 2≤y≤b+n/2.Rounding processing is done to the coordinate range acquired, and is calculated according to the coordinate range after rounding and needs the LED lighted mono- First (17) position No.: To be rounded symbol downwards；

When it is round for selecting LED light source shape, LED unit (17) coordinate range for needing to light are as follows: (x-a)²+(y-b)²≤ r².Rounding processing is done to the coordinate range acquired, and calculates the LED unit (17) for needing to light according to the coordinate range after rounding Position No.:WhereinI ' and j ' It is the row number and column number of round domain center LED unit (17) respectively；

When selecting LED light source shape is annular, LED unit (17) coordinate range for needing to light are as follows: r1²≤(x-a)²+(y-b)²≤ r2².Rounding processing is done to the coordinate range acquired, and calculates the LED unit (17) for needing to light according to the coordinate range after rounding Position No.:Wherein I ' and j ' is the row number and column number of annulus center LED unit (17) respectively；

LED unit (17) brightness value for needing to light is calculated, detailed process is as follows:

Assuming that LED unit (17) brightness value is lightness, setting center LED unit (17) brightness is 127, edge LED unit (17) brightness is 255；Remaining LED unit (17) brightness is using the distance away from center LED unit (17) as standard, in [127,255] Linear value is carried out in integer range；

When selecting LED light source shape for rectangle, position coordinates are LED unit (17) brightness value of (x, y) are as follows:Wherein max { ... } is the maximum value taken in bracket,To take downwards Integral symbol；

When it is round for selecting LED light source shape, position coordinates are LED unit (17) brightness value of (x, y) are as follows:

When it is annular for selecting LED light source shape, position coordinates are LED unit (17) brightness value of (x, y) are as follows:

After LED unit (17) parameter information calculates, one four one-dimension array comdata [4], data lattice are generated Formula is integer int, and carries out assignment processing to the array, is enabled

Comdata [0]=lightness indicates the brightness value of LED unit (17)；

Comdata [1]=delay indicates LED unit (17) illumination interval；

Comdata [2]=i indicates LED unit (17) row number；

Comdata [3]=j indicates LED unit (17) column number；

Wherein delay is LED unit (17) the lighting hours interval being artificially arranged；

Numerical value array comdata [4] is sequentially written in computer (1) according to set serial communication frequency after assignment Serial communication interface simultaneously saves, and slave computer is waited to read.

11. the microscopic imaging device according to claim 7 adaptively illuminated based on high-power LED lighteness carries out micro- Imaging measurement method, it is characterised in that in step 1.3):

Row port array RowPin [M] and column port array ColPin [N] are generated first in single-chip microcontroller, wherein M is LED array (13) the row number of LED unit (17) in, N are the port number of decoding relay (15) signal input part.Then row port number is given Group carries out assignment with column port array, with the mapping one by one established between ranks signal and microcontroller development board (8) port I/O, RowPin [M]={ ... } is enabled, RowPin [0] to RowPin [M] is followed successively by microcontroller development board (8) simulation output interface program Number；ColPin [N]={ ... } is enabled, ColPin [0] to ColPin [N] is followed successively by microcontroller development board (8) digital output port Program number；

When it is true value true that single-chip microcontroller, which receives the serial communication state from computer (1), the array of serial ports transmission is read Comdata [4], and fractionation reading is carried out to the array, identification obtains brightness lightness, the illumination frequencies of LED unit (17) F and position No. (i, j).Meanwhile row signal array row [M] and column signal array column [N] are generated in single-chip microcontroller, In the ignition period of one LED unit (17), assignment input is carried out to two signal arrays；

Row signal assignment: the simulation output port that program number is RowPin [i] is assigned a value of lightness, even row [RowPin [i]]=lightness, the other elements of row signal array row [M] are assigned a value of 0；

Column signal assignment: it is converted by system and decimal system column number j is converted into N bit binary number, and be assigned to columns group Column [N], even column [N]=j₍₂₎, wherein j₍₂₎For the position the N binary representation of decimal number j.Then successively by program The digital output port that number is ColPin [N] is assigned a value of column [N]；

The setting assignment waiting time is delay, is enabledWithin the waiting time, slave computer passes through access microcomputer development The port I/O of plate (8) carries out data read operation, after the waiting time, terminates the assignment operation in this stage, entrance is next The ignition period of LED unit (17).

12. the microscopic imaging device according to claim 7 adaptively illuminated based on high-power LED lighteness carries out micro- Imaging measurement method, it is characterised in that in step 1.4), column selection it is logical with row gating the specific implementation process is as follows:

Column selection is logical: after column selection messenger j is calculated in decoding relay (15), arranging electricity by the selected jth of relay control Road conducting, rest channels shutdown, only jth column LED unit (17) has electric current input, that is, it is logical to realize column selection；

Row gating: after the PWM input terminal of LED drive chip (14) receives the row gating signal from microcontroller development board (8), Parameter is inputted according to each road PWM, adjusts the size of current in each channel in load feedback end, not selected channel circuit is turned off, The i-th row current value being strobed then is adjusted toWherein V_LEDFor the conduction voltage drop of LED unit (17), V_outFor switch power source output voltage, R_iFor load resistance (16) resistance value of the i-th row.

13. the microscopic imaging device according to claim 8 adaptively illuminated based on high-power LED lighteness carries out micro- Imaging measurement method, it is characterised in that in step 3.1), the initial guess of microcobjective (5) pupil function P (u, v) are as follows:

Wherein u and v is light field frequency domain coordinates；f₀For the cutoff frequency of microcobjective (5).

14. the microscopic imaging device according to claim 8 adaptively illuminated based on high-power LED lighteness carries out micro- Imaging measurement method, it is characterised in that in step 3.2), in LED unit (17) in each submodule state corresponding sample frequency spectrum aperture Heart coordinateCalculation formula it is as follows:

Wherein x and y is respectively the plane abscissa and plane ordinate of the i-th row jth column LED unit (17)；Δx_sWith Δ y_sRespectively For the lateral distance and fore-and-aft distance of s-th submodule state and LED unit (17) center；H indicates LED array (3) and sample room Vertical range；λ indicates LED illumination optical wavelength.

15. the microscopic imaging device according to claim 8 adaptively illuminated based on high-power LED lighteness carries out micro- Imaging measurement method, it is characterised in that in step 3.3), the corresponding sample low resolution frequency spectrum of s-th of submodule state isCalculation formula are as follows:

Wherein subscript k indicates kth time iteration；(i, j) indicates the ranks coordinate of the LED unit (17)；S indicates s-th of submodule state；Indicate the high resolution spectrum sub-aperture of sample；

Then inverse Fourier transform is made to low resolution frequency spectrum, obtains the corresponding low resolution image planes complex amplitude of each submodule stateI.e. Indicate inverse Fourier transform.

16. the microscopic imaging device according to claim 8 adaptively illuminated based on high-power LED lighteness carries out micro- Imaging measurement method, it is characterised in that in step 3.4), the corresponding image planes complex amplitude more new formula of each submodule state is as follows:

The light intensity total value of imaging plane when calculating each mode first while illuminatingCalculation formula are as follows:

In formula | ... | two-dimensional complex number matrix norm is sought in expression；

It is then based on the ratio of each mode corresponding image planes intensity value and light intensity total value, is recorded using optical detector (7) Low resolution intensity image is updated the low resolution image planes complex amplitude of each mode, calculation formula are as follows:

WhereinFor the updated low resolution image planes complex amplitude of each mode,It is mono- for the i-th row jth column LED The intensity image that optical detector (7) is recorded when first (17) illumination sample.

17. the microscopic imaging device according to claim 8 adaptively illuminated based on high-power LED lighteness carries out micro- Imaging measurement method, it is characterised in that in step 3.5), image planes complex amplitude updates low resolution corresponding to each mode in front and back Spectrum differenceCalculation formula are as follows:

In formulaFourier transformation is made in expression；

The calculation formula that the frequency spectrum sub-aperture in the corresponding high resolution spectrum of each mode is updated based on the spectrum difference Are as follows:

In formulaIndicate updated sample high resolution spectrum sub-aperture；The function is sought in subscript * expression Conjugate function；|...|_maxThe maximum value of two-dimensional complex number matrix norm is sought in expression,Indicate sample more High resolution spectrum sub-aperture before new；

The more new formula of microcobjective (5) pupil function are as follows:

P ' in formula^k(u, v) indicates updated microcobjective (5) pupil function.