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In this section, simulation experiments are carried out on the collected image sequence with focus and without focus respectively.
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Firstly, the simulated experimental samples are selected. The intensity and phase images of the focus plane are shown in Fig.5(a) and Fig.5(b) respectively. The simulated image size is set to 256×256 pixel. Setting the focus position to 0 position. Firstly, five defocus images are obtained respectively before and after the focus plane by angular spectrum propagation, as shown in Fig.5(c). EDR-TIE algorithm is used directly for 11 intensity images which include focus image. The edge duty ratio detection results are shown in Fig.5(d), and it can be found that the focus image located at 0 position. The phase solution is carried out by the intensity image and the corresponding defocus images where the 0 position is located, and the result is shown in Fig.5(d).
Figure 5. Simulation experiment. (a) Set intensity image; (b) Set phase image; (c) A series of intensity images (including focus) of angular spectrum propagation; (d) EDR-TIE positioning calculation results and 0 position phase results graph; (e) AF-TIE positioning calculation result graph
The cycle operations continue to carry out by the AF-TIE algorithm. Using the intensity image of the 0 position and the phase image obtained by the solution to do angular spectrum propagate to the in-focus side, thus 11 intensity images are obtained, then the edge duty ratio detection is carried out again. If it found that the position is consistent with which obtain by EDR-TIE algorithm, as shown in Fig.5(e). Therefore, the 0 postion is considered as the optimal focus position which matches the set focus position.
Assuming that the original phase is
$\phi $ , the retrieved phase is${\phi _r}$ . Define error formula as[15]:The error of the phase result between the optimal focus position and the set position is 0.3050. It can be seemed that both EDR-TIE and AF-TIE algorithm proposed in this paper can locate the optimal focus position when the focus image is included.
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The intensity and phase in the focus plane of the simulated experimental sample are shown in Fig.6(a) and Fig.6(b) respectively. The distance between the two adjacent images is 100 μm, 0 position is the real focus position, and defocus images at positions from −8 to −13 are obtained by angular spectrum propagation, as shown in Fig.6(c). There is no focus image in this series of images. EDR-TIE algorithm is directly calculated from these 6 images. The positioning result is shown in Fig.6(d), and it can be found that it is located at the position of −8. We use angle spectrum by images at −8 position to obtain the intensity at −7 position. Then, using the intensity images at positions −9, −8, and −7 to obtain the phase of −8 position through TIE and the result is shown in Fig.6(e). Obviously, the EDR-TIE positioning result is inaccurate. If we want to obtain the focus image further, we usually need to collect a large number of intensity images again. Using AF-TIE algorithm in this paper, unidirectional angular spectrum propagation and edge duty ratio detection are carried out from the intensity image and the phase image at −8 position obtained by solution, and it is found that the focus plane is located at 0 position. The result is shown in Fig.6(f). Then move the translation table to collect the intensity image and the corresponding defocus images in 0 position, and the phase result show in Fig.6(f). Once again, one-directon angular spectrum propagation and edge duty ratio calculation are carried out to obtain the result in Fig.6(g), and it is found that it is located at its own position. Therefore, the 0 position is the optimal focus position, which is also the preseted. Then ending the cycle. This process reduce the collection number of intensity images.
Figure 6. Simulates experiment. (a) Set intensity image; (b) Set phase image; (c) A series of intensity images (unfocused) of angular spectrum propagation; (d) EDR-TIE positioning calculation results; (e) Phase retrieval results of EDR-TIE positioning positions; (f) AF-TIE positioning calculation results and phase retrieval results; (g) A series of intensity image AF-TIE positioning calculation results propagated by the 0 position angular spectrum in Fig.6(f)
In order to further compare the phase results of EDR-TIE algorithm and AF-TIE algorithm, an image quality evaluation index SSIM (structural similarity index) is introduced to measure the image structural similarity[16].
With the two images
$X$ and$Y$ entered,${\mu _X}$ and${\mu _Y}$ represent the average of$X$ and$Y$ ,${\sigma _X}$ and${\sigma _Y}$ represent the standard deviation of$X$ and$Y$ ,${\sigma _{XY}}$ is the covariance of$X$ and$Y$ ,${C_1}$ and${C_2}$ are the constants.The error and similarity between the phase results of EDR-TIE algorithm and AF-TIE algorithm and the set phase are calculated by Eq.(13) and Eq.(14) respectively, as shown in Tab.1. The effectiveness and accuracy of the algorithm proposed in this paper are proved.
Algorithm
evaluation indexRMSE SSIM EDR-TIE 0.4690 0.9687 AF-TIE 0.3050 0.9866 Table 1. Evaluation results of different algorithms
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In this section, real experiments are carried out on samples of continuous distribution and discrete distribution respectively. A small amount of intensity images are collected by inverted microscope (MI52, Mshot), and the acquisition interval of adjacent intensity images is 2 μm. The experimental device and internal optical path are shown in Fig.7. The illumination light source is the LED white light in the microscope. Firstly, it passes through a optical filter with a center wavelength of 532 nm and half-peak bandwidth of 22 nm. Then the light intensity adjusted by a light collector and an aperture diaphragm. Next, we obtain uniform collimated light through a light collector. The light irradiates the sample through the objective lens, the reflector and the lens barrel lens. Finally, the enlarged object light field is transmited to the camera port of the microscope. The resolution of CCD (MS60, Mshot) used in the experiment is 2048×2048 pixel.
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The continuous distributed cross-cutting cells of plant rhizomes are selected as sample in the real experiment of continuous distributed samples. EDR-TIE algorithm is used to locate the collected intensity images. The results are shown in Fig.8(a), and the positioning results are ideally located at the head or tail of a series of images. The phase is solved by using the intensity image and the corresponding defocus images of EDR-TIE positioning position, as shown in Fig.8(b). 10 intensity images are obtained from the EDR-TIE positioning position through angle spectrum propagated, since a new series of intensity images are obtained. Then AF-TIE calculation and edge duty ratio positioning are carried out. The positioning position is shown in Fig.8(d). Move the translation table to the AF-TIE positioning position to obtain the corresponding intensity images, then solve the phase, as shown in Fig.8(e). Once again, 10 intensity images are obtained through forward angle spectrum propagation, and edge duty ratio positioning is carried out. As shown in Fig.8(g). It is found that the located position has not changed. So the cycle can be ended.
Figure 8. Real experiment-plant rhizome cross section. (a) EDR-TIE positioning calculation result diagram; (b) EDR-TIE positioning position phase result diagram; (c) Partially enlarged phase result diagram of (b); (d) AF-TIE positioning calculation result diagram; (e) AF-TIE positioning position phase result diagram; (f) Partially enlarged phase result diagram of (e); (g) Optimal focus position angle spectrum AF-TIE positioning calculation result diagram
Figure8(c) and Figure8(f) are local amplification images of the phase results of Figure8(b) and Figure8(e) respectively. It can be seen that the optimal focus position can be located through several cycles by using AF-TIE algorithm.
In order to quantitatively evaluate the clarity of the retrieval phase, we use the edge duty ratio to compare the retrieval phase of EDR-TIE algorithm with AF-TIE algorithm, and the results are shown in Tab.2. It can be seen from the comparison result that when the initial image sequence does not contain focus image, the edge duty ratio of retrieval phase of the AF-TIE positioning position is smaller than the EDR-TIE positioning position, that is, AF-TIE algorithm has higher accuracy than EDR-TIE algorithm.
Focus method EDR-TIE AF-TIE Edge duty ratio 0.142 0.138 Table 2. Evaluation of retrieval phase by different algorithms
In order to further prove that the focus position located by AF-TIE algorithm is the optimal focus position. We move the translation stage of the microscope to the position of 7, which is the focus position located by AF-TIE algorithm, then taking 3 images at the same distance before and after the 7 position (the interval is 2 μm) and directly using the edge duty ratio to locate the the optimal focus image of the 7 images. The 7 images and the positioning results are shown in Fig.9(a) and Fig.9(b). It can be seen from the results that the real shot images are also located at position 7. That is, the positioning result of the edge duty ratio on the real shot image is consistent with the positioning result of AF-TIE, which proves the accuracy of the algorithm in this paper.
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In this experiment, microlense (Thorlab) was choosed as sample, and the lens material was fused Shi Ying with refractive index of 1.458. EDR-TIE positioning results are shown in Fig.10(a), then solve the corresponding phase and cycle. The best focus positioning results in this paper are shown in Fig.10(b). Move the translation table to the best positioning position and capture the corresponding intensity image, then solve the phase. The phase results of the best positioning positions of EDR-TIE and AF-TIE are shown in the Fig.10(c) and Fig.10(d) respectively. The true height of the microlens array is 1.11 μm. The height of the microlens array at EDR-TIE position is 1.2805 μm with error of 15.3%. The height of the microlens array at optimal focusing position of AF-TIE is 1.1739 μm with error of 5.7%. The results show that the optimal focal plane phase height obtained by AF-TIE algorithm in this paper is closer to the real value.
Figure 10. Actual experience micro lens array: (a) EDR-TIE positioning results; (b) AF-TIE was the best position result; (c) The optimal focus position angular spectrum AF-TIE positioning calculation result; (d) EDR-TIE position phase result and solid line section height curve; (e) AF-TIE position phase results and solid line section height curve
Phase retrieval based on the transport of intensity equation under adaptive focus
doi: 10.3788/IRLA20210231
- Received Date: 2021-12-10
- Rev Recd Date: 2022-01-20
- Publish Date: 2022-04-07
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Key words:
- phase retrieval /
- transport of intensity equation /
- adaptive focus /
- edge duty ratio
Abstract: The non-interference phase retrieval method based on the transport of intensity equation was a method to obtain the phase by solving the intensity images. In the process of image acquisition, the selection of in-focus image was very important. But it was usually determined by subjective methods, which led to inaccurate in-focus positioning, thus affecting the accuracy of phase results. Firstly, an phase retrieval method based on the transport of intensity equation under adaptive focus was proposed; Secondly, the edge duty ratio was used to locate the acquired images in this algorithm. After solving the phase, the optimal focus position was located when the edge duty ratio locating position kept unchanging by the circular angular spectrum propagation; Finally, the phase of the sample was solved by using the transport of intensity equation. The result show that this algorithm not only improved the accuracy of phase retrieval, but also reduced the time to obtain a large number of images. In the simulation experiment, the correlation coefficient between the retrieval phase and the original phase reached 0.9866, and the RMSE error is 0.3050. In the actual experiment of microlens array, the error between the true height of the microlens and the height solves by the phase retrieval method proposed is only 5.7%, which proves that the algorithm can locate the optimal focus position in the field of microscopic imaging. And the algorithm is conducive to the development of auto-focus technology and improves the accuracy of phase retrieval.