Research article

Cell segmentation in fluorescence microscopy images based on multi-scale histogram thresholding


  • Received: 18 May 2023 Revised: 07 August 2023 Accepted: 08 August 2023 Published: 14 August 2023
  • Cell segmentation from fluorescent microscopy images plays an important role in various applications, such as disease mechanism assessment and drug discovery research. Exiting segmentation methods often adopt image binarization as the first step, through which the foreground cell is separated from the background so that the subsequent processing steps can be greatly facilitated. To pursue this goal, a histogram thresholding can be performed on the input image, which first applies a Gaussian smoothing to suppress the jaggedness of the histogram curve and then exploits Rosin's method to determine a threshold for conducting image binarization. However, an inappropriate amount of smoothing could lead to the inaccurate segmentation of cells. To address this crucial problem, a multi-scale histogram thresholding (MHT) technique is proposed in the present paper, where the scale refers to the standard deviation of the Gaussian that determines the amount of smoothing. To be specific, the image histogram is smoothed at three chosen scales first, and then the smoothed histogram curves are fused to conduct image binarization via thresholding. To further improve the segmentation accuracy and overcome the difficulty of extracting overlapping cells, our proposed MHT technique is incorporated into a multi-scale cell segmentation framework, in which a region-based ellipse fitting technique is adopted to identify overlapping cells. Extensive experimental results obtained on benchmark datasets show that the new method can deliver superior performance compared to the current state-of-the-arts.

    Citation: Yating Fang, Baojiang Zhong. Cell segmentation in fluorescence microscopy images based on multi-scale histogram thresholding[J]. Mathematical Biosciences and Engineering, 2023, 20(9): 16259-16278. doi: 10.3934/mbe.2023726

    Related Papers:

  • Cell segmentation from fluorescent microscopy images plays an important role in various applications, such as disease mechanism assessment and drug discovery research. Exiting segmentation methods often adopt image binarization as the first step, through which the foreground cell is separated from the background so that the subsequent processing steps can be greatly facilitated. To pursue this goal, a histogram thresholding can be performed on the input image, which first applies a Gaussian smoothing to suppress the jaggedness of the histogram curve and then exploits Rosin's method to determine a threshold for conducting image binarization. However, an inappropriate amount of smoothing could lead to the inaccurate segmentation of cells. To address this crucial problem, a multi-scale histogram thresholding (MHT) technique is proposed in the present paper, where the scale refers to the standard deviation of the Gaussian that determines the amount of smoothing. To be specific, the image histogram is smoothed at three chosen scales first, and then the smoothed histogram curves are fused to conduct image binarization via thresholding. To further improve the segmentation accuracy and overcome the difficulty of extracting overlapping cells, our proposed MHT technique is incorporated into a multi-scale cell segmentation framework, in which a region-based ellipse fitting technique is adopted to identify overlapping cells. Extensive experimental results obtained on benchmark datasets show that the new method can deliver superior performance compared to the current state-of-the-arts.



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    [1] A. Gharipour, A. W. C. Liew, Segmentation of cell nuclei in fluorescence microscopy images: An integrated framework using level set segmentation and touching-cell splitting, Pattern Recognit., 58 (2016), 1–11. https://doi.org/10.1016/j.patcog.2016.03.030 doi: 10.1016/j.patcog.2016.03.030
    [2] D. Riccio, N. Brancati, M. Frucci, D. Gragnaniello, A new unsupervised approach for segmenting and counting cells in high-throughput microscopy image sets, IEEE J. Biomed. Health. Inf., 23 (2019), 437–448. https://doi.org/10.1109/JBHI.2018.2817485 doi: 10.1109/JBHI.2018.2817485
    [3] F. H. D. Araújo, R. R. V. Silva, F. N. S. Medeiros, J. F. R. Neto, H. C. P. Oliveira, Active contours for overlapping cervical cell segmentation, Int. J. Biomed. Eng. Technol., 35 (2021), 70–92. https://doi.org/10.1504/IJBET.2021.112834 doi: 10.1504/IJBET.2021.112834
    [4] Z. Wang, Z. Wang, A generic approach for cell segmentation based on Gabor filtering and area-constrained ultimate erosion, Artif. Intell. Med., 107 (2020), 101929. https://doi.org/10.1016/j.artmed.2020.101929 doi: 10.1016/j.artmed.2020.101929
    [5] K. Hajdowska, S. Student, D. Borys, Graph based method for cell segmentation and detection in live-cell fluorescence microscope imaging, Artif. Intell. Med., 71 (2022), 103071. https://doi.org/10.1016/j.bspc.2021.103071 doi: 10.1016/j.bspc.2021.103071
    [6] J. L. Mueller, Z. T. Harmany, J. K. Mito, S. A. Kennedy, Y. Kim, L. Dodd, et al., Quantitative segmentation of fluorescence microscopy images of heterogeneous tissue: Application to the detection of residual disease in tumor margins, PLoS One, 8 (2013), e66198. https://doi.org/10.1371/journal.pone.0066198 doi: 10.1371/journal.pone.0066198
    [7] M. Zhao, J. An, H. Li, J. Zhang, S. T. Li, X. M. Li, et al., Segmentation and classification of two-channel C. elegans nucleus-labeled fluorescence images, BMC Bioinf., 18 (2017), 1–13. https://doi.org/10.1186/s12859-017-1817-3 doi: 10.1186/s12859-017-1817-3
    [8] Z. Wang, Z. Wang, Robust cell segmentation based on gradient detection, Gabor filtering and morphological erosion, Biomed. Signal Process. Control, 65 (2021), 102390. https://doi.org/10.1016/j.bspc.2020.102390 doi: 10.1016/j.bspc.2020.102390
    [9] M. Salvi, U. Morbiducci, F. Amadeo, R. Santoro, F. Angelini, I. Chimenti, et al., Automated segmentation of fluorescence microscopy images for 3D cell detection in human-derived cardiospheres, Sci. Rep., 9 (2019), 6644. https://doi.org/10.1038/s41598-019-43137-2 doi: 10.1038/s41598-019-43137-2
    [10] D. Jia, C. Zhang, N. Wu, Z. Guo, H. Ge, Multi-layer segmentation framework for cell nuclei using improved GVF Snake model, Watershed, and ellipse fitting, Biomed. Signal Process. Control, 67 (2021), 102516. https://doi.org/10.1016/j.bspc.2021.102516 doi: 10.1016/j.bspc.2021.102516
    [11] T. Vicar, J. Balvan, J. Jaros, F. Jug, R. Kolar, M. Masarik, et al., Cell segmentation methods for label-free contrast microscopy: Review and comprehensive comparison, BMC Bioinf., 20 (2019), 1–25. https://doi.org/10.1186/s12859-019-2880-8 doi: 10.1186/s12859-019-2880-8
    [12] T. M. S. Mulyana, Herlina, Evenly brightening using kurtosis gaussian pattern to simplify image binarization, in Journal of Physics: Conference Series, 1397 (2019), 012076. https://doi.org/10.1088/1742-6596/1397/1/012076
    [13] P. D. Raju, G. Neelima, Image segmentation by using histogram thresholding, Int. J. Comput. Sci. Eng. Technol., 2 (2012), 776–779.
    [14] S. Ram, J. J. Rodriguez, Size-invariant detection of cell nuclei in microscopy images, IEEE Trans. Med. Imaging, 35 (2016), 1753–1764. https://doi.org/10.1109/TMI.2016.2527740 doi: 10.1109/TMI.2016.2527740
    [15] A. A. Ewees, M. A. Elaziz, M. A. A. Al-Qaness, H. A. Khalil, S. Kim, Improved artificial bee colony using sine-cosine algorithm for multi-level thresholding image segmentation, IEEE Access, 8 (2020), 26304–26315. https://doi.org/10.1109/ACCESS.2020.2971249 doi: 10.1109/ACCESS.2020.2971249
    [16] X. Yang, R. Wang, D. Zhao, F. Yu, A. A. Heidari, Z. Xu, et al., Multi-level threshold segmentation framework for breast cancer images using enhanced differential evolution, Biomed. Signal Process. Control, 80 (2023), 104373. https://doi.org/10.1016/j.bspc.2022.104373 doi: 10.1016/j.bspc.2022.104373
    [17] S. E. A. Raza, L. Cheung, D. Epstein, S. Pelengaris, M. Khan, N. M. Rajpoot, Mimo-net: A multi-input multi-output convolutional neural network for cell segmentation in fluorescence microscopy images, in 2017 IEEE 14th International Symposium on Biomedical Imaging (ISBI 2017), (2017), 337–340. https://doi.org/10.1109/ISBI.2017.7950532
    [18] H. Wang, P. Cao, J. Wang, O. R. Zaiane, Uctransnet: Rethinking the skip connections in u-net from a channel-wise perspective with transformer, in Proceedings of the AAAI Conference on Artificial Intelligence, 36 (2022), 2441–2449. https://doi.org/10.1609/aaai.v36i3.20144
    [19] H. Cao, Y. Wang, J. Chen, D. Jiang, X. Zhang, Q. Tian, et al., Swin-unet: Unet-like pure transformer for medical image segmentation, in European Conference on Computer Vision, (2023), 205–218. https://doi.org/10.1007/978-3-031-25066-8_9
    [20] D. K. Prasad, M. K. H. Leung, C. Quek, Ellifit: An unconstrained, non-iterative, least squares based geometric ellipse fitting method, Pattern Recognit., 46 (2013), 1449–1465. https://doi.org/10.1016/j.patcog.2012.11.007 doi: 10.1016/j.patcog.2012.11.007
    [21] Z. Shen, M. Zhao, X. Jia, Y. Liang, L. Fan, D. M. Yan, Combining convex hull and directed graph for fast and accurate ellipse detection, Graphical Models, 116 (2021), 101110. https://doi.org/10.1016/j.gmod.2021.101110 doi: 10.1016/j.gmod.2021.101110
    [22] S. Zafari, T. Eerola, J. Sampo, H. Kälviäinen, H. Haario, Segmentation of overlapping elliptical objects in silhouette images, IEEE Trans. Image Process., 24 (2015), 5942–5952. https://doi.org/10.1109/TIP.2015.2492828 doi: 10.1109/TIP.2015.2492828
    [23] P. K. Das, S. Meher, R. Panda, A. Abraham, An efficient blood-cell segmentation for the detection of hematological disorders, IEEE Trans. Cybern., 52 (2022), 10615–10626. https://doi.org/10.1109/TCYB.2021.3062152 doi: 10.1109/TCYB.2021.3062152
    [24] Y. Al-Kofahi, W. Lassoued, W. Lee, B. Roysam, Improved automatic detection and segmentation of cell nuclei in histopathology images, IEEE Trans. Biomed. Eng., 57 (2009), 841–852. https://doi.org/10.1109/TBME.2009.2035102 doi: 10.1109/TBME.2009.2035102
    [25] P. Rosin, Unimodal thresholding, Pattern Recognit., 34 (2001), 2083–2096. https://doi.org/10.1016/S0031-3203(00)00136-9
    [26] J. Liu, J. Xie, B. Li, B. Hu, Regularized cubic B-spline collocation method with modified L-curve criterion for impact force identification, IEEE Access, 8 (2020), 36337–36349. http://doi.org/10.1109/ACCESS.2020.2973919 doi: 10.1109/ACCESS.2020.2973919
    [27] J. Antoni, J. Idier, S. Bourguignon, A bayesian interpretation of the L-curve, Inverse Probl., 39 (2023), 065016. http://doi.org/10.1088/1361-6420/accdfc doi: 10.1088/1361-6420/accdfc
    [28] W. V. Drongelen, Signal Processing for Neuroscientists, 2nd edition, Academic Press, 2018. https://doi.org/10.1016/B978-0-12-370867-0.X5000-1
    [29] C. Panagiotakis, A. Argyros, Region-based fitting of overlapping ellipses and its application to cells segmentation, Image Vision Comput., 93 (2020), 103810. https://doi.org/10.1016/j.imavis.2019.09.001 doi: 10.1016/j.imavis.2019.09.001
    [30] M. Liao, Y. Q. Zhao, X. H. Li, P. S. Dai, X. W. Xu, J. Zhang, et al., Automatic segmentation for cell images based on bottleneck detection and ellipse fitting, Neurocomputing, 173 (2016), 615–622. https://doi.org/10.1016/j.neucom.2015.08.006 doi: 10.1016/j.neucom.2015.08.006
    [31] C. Panagiotakis, A. Argyros, Parameter-free modelling of 2D shapes with ellipses, Pattern Recognit., 53 (2016), 259–275. https://doi.org/10.1016/j.patcog.2015.11.004 doi: 10.1016/j.patcog.2015.11.004
    [32] B. Zhong, K. K. Ma, On the convergence of planar curves under smoothing, IEEE Trans. Image Process., 19 (2010), 2171–2189. https://doi.org/10.1109/TIP.2010.2046807 doi: 10.1109/TIP.2010.2046807
    [33] L. P. Coelho, A. Shariff, R. F. Murphy, Nuclear segmentation in microscope cell images: A hand-segmented dataset and comparison of algorithms, in 2009 IEEE International Symposium on Biomedical Imaging: From Nano to Macro, (2009), 518–521. https://doi.org/10.1109/ISBI.2009.5193098
    [34] N. Kumar, R. Verma, S. Sharma, S. Bhargava, A. Vahadane, A. Sethi, A dataset and a technique for generalized nuclear segmentation for computational pathology, IEEE Trans. Med. Imaging, 36 (2017), 1550–1560. https://doi.org/10.1109/TMI.2017.2677499 doi: 10.1109/TMI.2017.2677499
    [35] J. P. Bergeest, K. Rohr, Efficient globally optimal segmentation of cells in fluorescence microscopy images using level sets and convex energy functionals, Med. Image Anal., 16 (2012), 1436–1444. https://doi.org/10.1016/j.media.2012.05.012 doi: 10.1016/j.media.2012.05.012
    [36] Y. T. Chen, A level set method based on the bayesian risk for medical image segmentation, Pattern Recognit., 43 (2010), 3699–3711. https://doi.org/10.1016/j.patcog.2010.05.027 doi: 10.1016/j.patcog.2010.05.027
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