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| import numpy as np | |
| from scipy.stats import pearsonr | |
| from .._shared.utils import check_shape_equality, as_binary_ndarray | |
| __all__ = [ | |
| 'pearson_corr_coeff', | |
| 'manders_coloc_coeff', | |
| 'manders_overlap_coeff', | |
| 'intersection_coeff', | |
| ] | |
| def pearson_corr_coeff(image0, image1, mask=None): | |
| r"""Calculate Pearson's Correlation Coefficient between pixel intensities | |
| in channels. | |
| Parameters | |
| ---------- | |
| image0 : (M, N) ndarray | |
| Image of channel A. | |
| image1 : (M, N) ndarray | |
| Image of channel 2 to be correlated with channel B. | |
| Must have same dimensions as `image0`. | |
| mask : (M, N) ndarray of dtype bool, optional | |
| Only `image0` and `image1` pixels within this region of interest mask | |
| are included in the calculation. Must have same dimensions as `image0`. | |
| Returns | |
| ------- | |
| pcc : float | |
| Pearson's correlation coefficient of the pixel intensities between | |
| the two images, within the mask if provided. | |
| p-value : float | |
| Two-tailed p-value. | |
| Notes | |
| ----- | |
| Pearson's Correlation Coefficient (PCC) measures the linear correlation | |
| between the pixel intensities of the two images. Its value ranges from -1 | |
| for perfect linear anti-correlation to +1 for perfect linear correlation. | |
| The calculation of the p-value assumes that the intensities of pixels in | |
| each input image are normally distributed. | |
| Scipy's implementation of Pearson's correlation coefficient is used. Please | |
| refer to it for further information and caveats [1]_. | |
| .. math:: | |
| r = \frac{\sum (A_i - m_A_i) (B_i - m_B_i)} | |
| {\sqrt{\sum (A_i - m_A_i)^2 \sum (B_i - m_B_i)^2}} | |
| where | |
| :math:`A_i` is the value of the :math:`i^{th}` pixel in `image0` | |
| :math:`B_i` is the value of the :math:`i^{th}` pixel in `image1`, | |
| :math:`m_A_i` is the mean of the pixel values in `image0` | |
| :math:`m_B_i` is the mean of the pixel values in `image1` | |
| A low PCC value does not necessarily mean that there is no correlation | |
| between the two channel intensities, just that there is no linear | |
| correlation. You may wish to plot the pixel intensities of each of the two | |
| channels in a 2D scatterplot and use Spearman's rank correlation if a | |
| non-linear correlation is visually identified [2]_. Also consider if you | |
| are interested in correlation or co-occurence, in which case a method | |
| involving segmentation masks (e.g. MCC or intersection coefficient) may be | |
| more suitable [3]_ [4]_. | |
| Providing the mask of only relevant sections of the image (e.g., cells, or | |
| particular cellular compartments) and removing noise is important as the | |
| PCC is sensitive to these measures [3]_ [4]_. | |
| References | |
| ---------- | |
| .. [1] https://docs.scipy.org/doc/scipy/reference/generated/scipy.stats.pearsonr.html | |
| .. [2] https://docs.scipy.org/doc/scipy/reference/generated/scipy.stats.spearmanr.html | |
| .. [3] Dunn, K. W., Kamocka, M. M., & McDonald, J. H. (2011). A practical | |
| guide to evaluating colocalization in biological microscopy. | |
| American journal of physiology. Cell physiology, 300(4), C723–C742. | |
| https://doi.org/10.1152/ajpcell.00462.2010 | |
| .. [4] Bolte, S. and Cordelières, F.P. (2006), A guided tour into | |
| subcellular colocalization analysis in light microscopy. Journal of | |
| Microscopy, 224: 213-232. | |
| https://doi.org/10.1111/j.1365-2818.2006.01706.x | |
| """ | |
| image0 = np.asarray(image0) | |
| image1 = np.asarray(image1) | |
| if mask is not None: | |
| mask = as_binary_ndarray(mask, variable_name="mask") | |
| check_shape_equality(image0, image1, mask) | |
| image0 = image0[mask] | |
| image1 = image1[mask] | |
| else: | |
| check_shape_equality(image0, image1) | |
| # scipy pearsonr function only takes flattened arrays | |
| image0 = image0.reshape(-1) | |
| image1 = image1.reshape(-1) | |
| return tuple(float(v) for v in pearsonr(image0, image1)) | |
| def manders_coloc_coeff(image0, image1_mask, mask=None): | |
| r"""Manders' colocalization coefficient between two image channels. | |
| Parameters | |
| ---------- | |
| image0 : (M, N) ndarray | |
| Input image (first channel). All pixel values should be non-negative. | |
| image1_mask : (M, N) ndarray of dtype bool | |
| Binary image giving the regions of interest in the second channel. | |
| Must have same shape as `image0`. | |
| mask : (M, N) ndarray of dtype bool, optional | |
| Only `image0` pixel values within `mask` are included in the calculation. | |
| Must have same shape as `image0`. | |
| Returns | |
| ------- | |
| mcc : float | |
| Manders' colocalization coefficient. | |
| Notes | |
| ----- | |
| Manders' colocalization coefficient (MCC) was developed in the context of | |
| confocal biological microscopy, to measure the fraction of colocalizing | |
| objects in each component of a dual-channel image. Out of the total | |
| intensity of, say, channel A, how much is found within the features | |
| (objects) of, say, channel B [1]_? The measure thus ranges from 0 for no | |
| colocalization to 1 for complete colocalization. | |
| MCC is commonly used to measure the colocalization of a particular protein | |
| in a subcelullar compartment. Typically, the mask for channel B is | |
| obtained by thresholding, to segment the features from the background. | |
| In this implementation, channel B is passed directly as a mask | |
| (`image1_mask`), leaving the segmentation step to the user (upstream). | |
| The implemented equation is: | |
| .. math:: | |
| mcc = \frac{\sum_i A_{i,coloc}}{\sum_i A_i} | |
| where | |
| - :math:`A_i` is the value of the :math:`i^{th}` pixel in `image0`, and | |
| - :math:`A_{i, coloc} = A_i B_i`, considering that :math:`B_i` is the | |
| (``True`` or ``False``) value of the :math:`i^{th}` pixel in | |
| `image1_mask` cast into int or float (``1`` or ``0``, respectively). | |
| MCC is sensitive to noise, with diffuse signal in the first channel | |
| inflating its value. Therefore, images should be processed beforehand to | |
| remove out-of-focus and background light [2]_. | |
| References | |
| ---------- | |
| .. [1] Manders, E.M.M., Verbeek, F.J. and Aten, J.A. (1993), Measurement of | |
| co-localization of objects in dual-colour confocal images. Journal | |
| of Microscopy, 169: 375-382. | |
| https://doi.org/10.1111/j.1365-2818.1993.tb03313.x | |
| https://imagej.net/media/manders.pdf | |
| .. [2] Dunn, K. W., Kamocka, M. M., & McDonald, J. H. (2011). A practical | |
| guide to evaluating colocalization in biological microscopy. | |
| American journal of physiology. Cell physiology, 300(4), C723–C742. | |
| https://doi.org/10.1152/ajpcell.00462.2010 | |
| """ | |
| image0 = np.asarray(image0) | |
| image1_mask = as_binary_ndarray(image1_mask, variable_name="image1_mask") | |
| if mask is not None: | |
| mask = as_binary_ndarray(mask, variable_name="mask") | |
| check_shape_equality(image0, image1_mask, mask) | |
| image0 = image0[mask] | |
| image1_mask = image1_mask[mask] | |
| else: | |
| check_shape_equality(image0, image1_mask) | |
| # check non-negative image | |
| if image0.min() < 0: | |
| raise ValueError("image contains negative values") | |
| sum = np.sum(image0) | |
| if sum == 0: | |
| return 0 | |
| return np.sum(image0 * image1_mask) / sum | |
| def manders_overlap_coeff(image0, image1, mask=None): | |
| r"""Manders' overlap coefficient | |
| Parameters | |
| ---------- | |
| image0 : (M, N) ndarray | |
| Image of channel A. All pixel values should be non-negative. | |
| image1 : (M, N) ndarray | |
| Image of channel B. All pixel values should be non-negative. | |
| Must have same dimensions as `image0` | |
| mask : (M, N) ndarray of dtype bool, optional | |
| Only `image0` and `image1` pixel values within this region of interest | |
| mask are included in the calculation. | |
| Must have ♣same dimensions as `image0`. | |
| Returns | |
| ------- | |
| moc: float | |
| Manders' Overlap Coefficient of pixel intensities between the two | |
| images. | |
| Notes | |
| ----- | |
| Manders' Overlap Coefficient (MOC) is given by the equation [1]_: | |
| .. math:: | |
| r = \frac{\sum A_i B_i}{\sqrt{\sum A_i^2 \sum B_i^2}} | |
| where | |
| :math:`A_i` is the value of the :math:`i^{th}` pixel in `image0` | |
| :math:`B_i` is the value of the :math:`i^{th}` pixel in `image1` | |
| It ranges between 0 for no colocalization and 1 for complete colocalization | |
| of all pixels. | |
| MOC does not take into account pixel intensities, just the fraction of | |
| pixels that have positive values for both channels[2]_ [3]_. Its usefulness | |
| has been criticized as it changes in response to differences in both | |
| co-occurence and correlation and so a particular MOC value could indicate | |
| a wide range of colocalization patterns [4]_ [5]_. | |
| References | |
| ---------- | |
| .. [1] Manders, E.M.M., Verbeek, F.J. and Aten, J.A. (1993), Measurement of | |
| co-localization of objects in dual-colour confocal images. Journal | |
| of Microscopy, 169: 375-382. | |
| https://doi.org/10.1111/j.1365-2818.1993.tb03313.x | |
| https://imagej.net/media/manders.pdf | |
| .. [2] Dunn, K. W., Kamocka, M. M., & McDonald, J. H. (2011). A practical | |
| guide to evaluating colocalization in biological microscopy. | |
| American journal of physiology. Cell physiology, 300(4), C723–C742. | |
| https://doi.org/10.1152/ajpcell.00462.2010 | |
| .. [3] Bolte, S. and Cordelières, F.P. (2006), A guided tour into | |
| subcellular colocalization analysis in light microscopy. Journal of | |
| Microscopy, 224: 213-232. | |
| https://doi.org/10.1111/j.1365-2818.2006.01 | |
| .. [4] Adler J, Parmryd I. (2010), Quantifying colocalization by | |
| correlation: the Pearson correlation coefficient is | |
| superior to the Mander's overlap coefficient. Cytometry A. | |
| Aug;77(8):733-42.https://doi.org/10.1002/cyto.a.20896 | |
| .. [5] Adler, J, Parmryd, I. Quantifying colocalization: The case for | |
| discarding the Manders overlap coefficient. Cytometry. 2021; 99: | |
| 910– 920. https://doi.org/10.1002/cyto.a.24336 | |
| """ | |
| image0 = np.asarray(image0) | |
| image1 = np.asarray(image1) | |
| if mask is not None: | |
| mask = as_binary_ndarray(mask, variable_name="mask") | |
| check_shape_equality(image0, image1, mask) | |
| image0 = image0[mask] | |
| image1 = image1[mask] | |
| else: | |
| check_shape_equality(image0, image1) | |
| # check non-negative image | |
| if image0.min() < 0: | |
| raise ValueError("image0 contains negative values") | |
| if image1.min() < 0: | |
| raise ValueError("image1 contains negative values") | |
| denom = (np.sum(np.square(image0)) * (np.sum(np.square(image1)))) ** 0.5 | |
| return np.sum(np.multiply(image0, image1)) / denom | |
| def intersection_coeff(image0_mask, image1_mask, mask=None): | |
| r"""Fraction of a channel's segmented binary mask that overlaps with a | |
| second channel's segmented binary mask. | |
| Parameters | |
| ---------- | |
| image0_mask : (M, N) ndarray of dtype bool | |
| Image mask of channel A. | |
| image1_mask : (M, N) ndarray of dtype bool | |
| Image mask of channel B. | |
| Must have same dimensions as `image0_mask`. | |
| mask : (M, N) ndarray of dtype bool, optional | |
| Only `image0_mask` and `image1_mask` pixels within this region of | |
| interest | |
| mask are included in the calculation. | |
| Must have same dimensions as `image0_mask`. | |
| Returns | |
| ------- | |
| Intersection coefficient, float | |
| Fraction of `image0_mask` that overlaps with `image1_mask`. | |
| """ | |
| image0_mask = as_binary_ndarray(image0_mask, variable_name="image0_mask") | |
| image1_mask = as_binary_ndarray(image1_mask, variable_name="image1_mask") | |
| if mask is not None: | |
| mask = as_binary_ndarray(mask, variable_name="mask") | |
| check_shape_equality(image0_mask, image1_mask, mask) | |
| image0_mask = image0_mask[mask] | |
| image1_mask = image1_mask[mask] | |
| else: | |
| check_shape_equality(image0_mask, image1_mask) | |
| nonzero_image0 = np.count_nonzero(image0_mask) | |
| if nonzero_image0 == 0: | |
| return 0 | |
| nonzero_joint = np.count_nonzero(np.logical_and(image0_mask, image1_mask)) | |
| return nonzero_joint / nonzero_image0 | |