1. Usage#

[1]:

%load_ext autoreload
%autoreload 2

import matplotlib.pyplot as plt
import numpy as np
import scipy
import seaborn as sns
import skimage
import skimage.io
import tifffile
from pympler.asizeof import asizeof

from nima import nima, segmentation

cm = plt.cm.inferno_r

fp = "../../tests/data/1b_c16_15.tif"

channels = ["G", "R", "C"]
# dark = io.imread('/home/dati/GBM_persson/analyses/15.02.05_cal-GBM5-pBJclop/dark/dark-25_500.tif')
# flat = io.imread('/home/dati/GBM_persson/analyses/15.02.05_cal-GBM5-pBJclop/flat/flat-C-dark-37bis_500.tif')

[2]:

bg_params = segmentation.BgParams(kind="li_adaptive")
bg_params

[2]:

BgParams(kind='li_adaptive', perc=0.1, radius=10, adaptive_radius=None, arcsinh_perc=80, erosion_disk=0, clip=False)

[3]:

d_im, n_ch, times = nima.read_tiff(fp, channels)

(4, 512, 512)

[4]:

d_im_bg, bgs, ff = nima.d_bg(d_im, downscale=(2, 2), bg_params=bg_params)
bgs

[4]:

	G	R	C
0	462.00	249.75	278.00
1	463.50	253.00	280.25
2	464.50	254.75	282.00
3	461.25	252.50	278.00

[5]:

d_im_bg["G"].__sizeof__() / 1024**2

[5]:

8.000137329101562

[6]:

ff["C"][1][0]

[6]:

../_images/tutorials_usage_6_0.png

[7]:

nima.d_mask_label(
    d_im_bg, threshold_method="yen", min_size=2000, channels=channels, watershed=0
)
d_im_bg.keys()

[7]:

dict_keys(['G', 'R', 'C', 'mask', 'labels'])

[8]:

f = nima.d_show(d_im_bg)

../_images/tutorials_usage_8_0.png

[9]:

meas, pr = nima.d_meas_props(d_im_bg)
f = nima.d_plot_meas(bgs, meas, channels=channels)

../_images/tutorials_usage_9_0.png

[10]:

plt.subplot(1, 2, 1)
skimage.io.imshow(d_im_bg["r_cl"][2], vmin=0.0, vmax=1.1)
plt.subplot(1, 2, 2)
skimage.io.imshow(d_im_bg["r_pH"][2], vmin=7.3, vmax=10.7)

/tmp/ipykernel_2661/3384179216.py:2: FutureWarning: `imshow` is deprecated since version 0.25 and will be removed in version 0.27. Please use `matplotlib`, `napari`, etc. to visualize images.
  skimage.io.imshow(d_im_bg["r_cl"][2], vmin=0.0, vmax=1.1)
/home/runner/work/nima/nima/.venv/lib/python3.13/site-packages/skimage/io/_plugins/matplotlib_plugin.py:158: UserWarning: Float image out of standard range; displaying image with stretched contrast.
  lo, hi, cmap = _get_display_range(image)
/tmp/ipykernel_2661/3384179216.py:4: FutureWarning: `imshow` is deprecated since version 0.25 and will be removed in version 0.27. Please use `matplotlib`, `napari`, etc. to visualize images.
  skimage.io.imshow(d_im_bg["r_pH"][2], vmin=7.3, vmax=10.7)
/home/runner/work/nima/nima/.venv/lib/python3.13/site-packages/skimage/io/_plugins/matplotlib_plugin.py:158: UserWarning: Float image out of standard range; displaying image with stretched contrast.
  lo, hi, cmap = _get_display_range(image)

[10]:

<matplotlib.image.AxesImage at 0x7f9ff1731950>

../_images/tutorials_usage_10_2.png

1.1. dev#

[11]:

c0 = d_im["C"][0]
g0 = d_im["G"][0]
r0 = d_im["R"][0]
tifffile.imshow(1 / c0 / c0)

[11]:

(<Figure size 988.8x604.8 with 2 Axes>,
 <Axes: >,
 <matplotlib.image.AxesImage at 0x7f9ff13f5810>)

../_images/tutorials_usage_12_1.png

[12]:

bg_params = segmentation.BgParams(kind="li_adaptive", erosion_disk=0)
bg_params

[12]:

BgParams(kind='li_adaptive', perc=0.1, radius=10, adaptive_radius=None, arcsinh_perc=80, erosion_disk=0, clip=False)

[13]:

im = c0
im = d_im["C"][1]
bg_params.adaptive_radius = int(im.shape[1] / 2)
if bg_params.adaptive_radius % 2 == 0:  # sk >0.12.0 check for even value
    bg_params.adaptive_radius += 1
bg_params

[13]:

BgParams(kind='li_adaptive', perc=0.1, radius=10, adaptive_radius=257, arcsinh_perc=80, erosion_disk=0, clip=False)

[14]:

bg_params.clip = False
m, title, lim = segmentation._bg_li_adaptive(im, bg_params=bg_params)
title, lim

[14]:

('li_adaptive adaptive_radius=257', None)

# The second is about 30-40% faster %timeit np.ma.masked_array(im, ~m).mean() %timeit im[m].mean()

[15]:

scipy.stats.distributions.norm.fit(im[m])

[15]:

(np.float64(278.5552310855508), np.float64(41.05308082242734))

[16]:

sns.histplot(im[m], kde=True, stat="density", log_scale=(False, True))

[16]:

<Axes: ylabel='Density'>

../_images/tutorials_usage_18_1.png

[17]:

fig = plt.hist(im[m], histtype="step", bins=32, log=1)
fig = plt.hist(im[m], histtype="step", bins=32, log=1)

../_images/tutorials_usage_19_0.png

[18]:

segmentation.calculate_bg_iteratively(im, probplot=True)

289.55334184900715 23.81726825652639
356

[18]:

BgResult(bg=np.float64(289.55334184900715), sd=np.float64(23.81726825652639), iqr=array([273., 288., 305.]), figures=[<Figure size 1200x400 with 4 Axes>])

../_images/tutorials_usage_20_2.png

1.2. background, AF, target cells#

[19]:

fp_mdtest = "/home/dati/GBM_persson/data/15.02.05_cal-GBM5-pBJclop/ph633/1_20_40.tif"
fp_mdtest = "/home/dati/dt-clop3/data/ionofori assay pH6.8 22.05.05/Clop FLATxy Gexp20ms Rexp3ms.tf8"

[20]:

nima.read_tiff(fp_mdtest, ["G", "C", "R"])

---------------------------------------------------------------------------
FileNotFoundError                         Traceback (most recent call last)
/tmp/ipykernel_2661/2636126201.py in ?()
----> 1 nima.read_tiff(fp_mdtest, ["G", "C", "R"])

~/work/nima/nima/src/nima/nima.py in ?(fp, channels)
     70     (3, 4)
     71
     72     """
     73     n_channels = len(channels)
---> 74     with TiffFile(fp) as tif:
     75         im = tif.asarray()
     76         axes = tif.series[0].axes
     77     idx = axes.rfind("T")

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size, omexml, _multifile, _useframes, _parent, **is_flags)
   4240                 raise ValueError('invalid OME-XML')
   4241             self._omexml = omexml
   4242             self.is_ome = True
   4243
-> 4244         fh = FileHandle(file, mode=mode, name=name, offset=offset, size=size)
   4245         self._fh = fh
   4246         self._multifile = True if _multifile is None else bool(_multifile)
   4247         self._files = {fh.name: self}

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size)
  13350         self._offset = -1 if offset is None else offset
  13351         self._size = -1 if size is None else size
  13352         self._close = True
  13353         self._lock = NullContext()
> 13354         self.open()
  13355         assert self._fh is not None

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self)
  13369             if self._mode not in {'rb', 'r+b', 'wb', 'xb'}:
  13370                 raise ValueError(f'invalid mode {self._mode}')
  13371             self._file = os.path.realpath(self._file)
  13372             self._dir, self._name = os.path.split(self._file)
> 13373             self._fh = open(self._file, self._mode, encoding=None)
  13374             self._close = True
  13375             self._offset = max(0, self._offset)
  13376         elif isinstance(self._file, FileHandle):

FileNotFoundError: [Errno 2] No such file or directory: '/home/dati/dt-clop3/data/ionofori assay pH6.8 22.05.05/Clop FLATxy Gexp20ms Rexp3ms.tf8'

[21]:

dim, md = nima.read_tiffmd(fp_mdtest, channels)

---------------------------------------------------------------------------
FileNotFoundError                         Traceback (most recent call last)
/tmp/ipykernel_2661/3787057078.py in ?()
----> 1 dim, md = nima.read_tiffmd(fp_mdtest, channels)

~/work/nima/nima/src/nima/nima.py in ?(fp, channels)
   1072 def read_tiffmd(fp: Path, channels: Sequence[str]) -> tuple[Array, Metadata]:
   1073     """Read multichannel TIFF timelapse image."""
   1074     n_channels = len(channels)
-> 1075     rdr = TiffReader(fp)
   1076     dim = da.from_zarr(rdr.aszarr())  # type: ignore[no-untyped-call]
   1077     md = Metadata(rdr)
   1078     if md.size_c[0] % n_channels:

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size, omexml, _multifile, _useframes, _parent, **is_flags)
   4240                 raise ValueError('invalid OME-XML')
   4241             self._omexml = omexml
   4242             self.is_ome = True
   4243
-> 4244         fh = FileHandle(file, mode=mode, name=name, offset=offset, size=size)
   4245         self._fh = fh
   4246         self._multifile = True if _multifile is None else bool(_multifile)
   4247         self._files = {fh.name: self}

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size)
  13350         self._offset = -1 if offset is None else offset
  13351         self._size = -1 if size is None else size
  13352         self._close = True
  13353         self._lock = NullContext()
> 13354         self.open()
  13355         assert self._fh is not None

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self)
  13369             if self._mode not in {'rb', 'r+b', 'wb', 'xb'}:
  13370                 raise ValueError(f'invalid mode {self._mode}')
  13371             self._file = os.path.realpath(self._file)
  13372             self._dir, self._name = os.path.split(self._file)
> 13373             self._fh = open(self._file, self._mode, encoding=None)
  13374             self._close = True
  13375             self._offset = max(0, self._offset)
  13376         elif isinstance(self._file, FileHandle):

FileNotFoundError: [Errno 2] No such file or directory: '/home/dati/dt-clop3/data/ionofori assay pH6.8 22.05.05/Clop FLATxy Gexp20ms Rexp3ms.tf8'

[22]:

md.stage_position

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[22], line 1
----> 1 md.stage_position

NameError: name 'md' is not defined

[23]:

im = dim[:, 0].mean(axis=0).compute()
plt.imshow(im)
plt.colorbar()

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[23], line 1
----> 1 im = dim[:, 0].mean(axis=0).compute()
      2 plt.imshow(im)
      3 plt.colorbar()

NameError: name 'dim' is not defined

[24]:

flat_im = scipy.ndimage.gaussian_filter(im, 150)
plt.imshow(flat_im)
plt.colorbar()

[24]:

<matplotlib.colorbar.Colorbar at 0x7f9fef535310>

../_images/tutorials_usage_27_1.png

[25]:

plt.plot(flat_im[1060, :])

---------------------------------------------------------------------------
IndexError                                Traceback (most recent call last)
Cell In[25], line 1
----> 1 plt.plot(flat_im[1060, :])

IndexError: index 1060 is out of bounds for axis 0 with size 512

[26]:

fpm = "/home/dan/workspace/nima_io/tests/data/t4_1.tif"

[27]:

nima.read_tiffmd(fpm, ["G", "R2", "R", "C"])

---------------------------------------------------------------------------
FileNotFoundError                         Traceback (most recent call last)
/tmp/ipykernel_2661/3851534406.py in ?()
----> 1 nima.read_tiffmd(fpm, ["G", "R2", "R", "C"])

~/work/nima/nima/src/nima/nima.py in ?(fp, channels)
   1072 def read_tiffmd(fp: Path, channels: Sequence[str]) -> tuple[Array, Metadata]:
   1073     """Read multichannel TIFF timelapse image."""
   1074     n_channels = len(channels)
-> 1075     rdr = TiffReader(fp)
   1076     dim = da.from_zarr(rdr.aszarr())  # type: ignore[no-untyped-call]
   1077     md = Metadata(rdr)
   1078     if md.size_c[0] % n_channels:

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size, omexml, _multifile, _useframes, _parent, **is_flags)
   4240                 raise ValueError('invalid OME-XML')
   4241             self._omexml = omexml
   4242             self.is_ome = True
   4243
-> 4244         fh = FileHandle(file, mode=mode, name=name, offset=offset, size=size)
   4245         self._fh = fh
   4246         self._multifile = True if _multifile is None else bool(_multifile)
   4247         self._files = {fh.name: self}

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size)
  13350         self._offset = -1 if offset is None else offset
  13351         self._size = -1 if size is None else size
  13352         self._close = True
  13353         self._lock = NullContext()
> 13354         self.open()
  13355         assert self._fh is not None

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self)
  13369             if self._mode not in {'rb', 'r+b', 'wb', 'xb'}:
  13370                 raise ValueError(f'invalid mode {self._mode}')
  13371             self._file = os.path.realpath(self._file)
  13372             self._dir, self._name = os.path.split(self._file)
> 13373             self._fh = open(self._file, self._mode, encoding=None)
  13374             self._close = True
  13375             self._offset = max(0, self._offset)
  13376         elif isinstance(self._file, FileHandle):

FileNotFoundError: [Errno 2] No such file or directory: '/home/dan/workspace/nima_io/tests/data/t4_1.tif'

[28]:

nima.read_tiffmd(fpm, ["G", "R2", "R", "C"])

---------------------------------------------------------------------------
FileNotFoundError                         Traceback (most recent call last)
/tmp/ipykernel_2661/3851534406.py in ?()
----> 1 nima.read_tiffmd(fpm, ["G", "R2", "R", "C"])

~/work/nima/nima/src/nima/nima.py in ?(fp, channels)
   1072 def read_tiffmd(fp: Path, channels: Sequence[str]) -> tuple[Array, Metadata]:
   1073     """Read multichannel TIFF timelapse image."""
   1074     n_channels = len(channels)
-> 1075     rdr = TiffReader(fp)
   1076     dim = da.from_zarr(rdr.aszarr())  # type: ignore[no-untyped-call]
   1077     md = Metadata(rdr)
   1078     if md.size_c[0] % n_channels:

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size, omexml, _multifile, _useframes, _parent, **is_flags)
   4240                 raise ValueError('invalid OME-XML')
   4241             self._omexml = omexml
   4242             self.is_ome = True
   4243
-> 4244         fh = FileHandle(file, mode=mode, name=name, offset=offset, size=size)
   4245         self._fh = fh
   4246         self._multifile = True if _multifile is None else bool(_multifile)
   4247         self._files = {fh.name: self}

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size)
  13350         self._offset = -1 if offset is None else offset
  13351         self._size = -1 if size is None else size
  13352         self._close = True
  13353         self._lock = NullContext()
> 13354         self.open()
  13355         assert self._fh is not None

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self)
  13369             if self._mode not in {'rb', 'r+b', 'wb', 'xb'}:
  13370                 raise ValueError(f'invalid mode {self._mode}')
  13371             self._file = os.path.realpath(self._file)
  13372             self._dir, self._name = os.path.split(self._file)
> 13373             self._fh = open(self._file, self._mode, encoding=None)
  13374             self._close = True
  13375             self._offset = max(0, self._offset)
  13376         elif isinstance(self._file, FileHandle):

FileNotFoundError: [Errno 2] No such file or directory: '/home/dan/workspace/nima_io/tests/data/t4_1.tif'

[29]:

from nima import utils

[30]:

bgd = {}
for i, c in enumerate(channels):
    bgd[c] = [
        segmentation.calculate_bg_iteratively(dim[t, i].compute())
        for t in range(md.size_t[0])
    ]

bgd

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[30], line 5
      1 bgd = {}
      2 for i, c in enumerate(channels):
      3     bgd[c] = [
      4         segmentation.calculate_bg_iteratively(dim[t, i].compute())
----> 5         for t in range(md.size_t[0])
      6     ]
      8 bgd

NameError: name 'md' is not defined

[31]:

for t in range(md.size_t[0]):
    dim[t, 0] -= bgd[channels[0]][t].bg

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[31], line 1
----> 1 for t in range(md.size_t[0]):
      2     dim[t, 0] -= bgd[channels[0]][t].bg

NameError: name 'md' is not defined

[32]:

plt.imshow(dim[0, 0])
plt.colorbar()

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[32], line 1
----> 1 plt.imshow(dim[0, 0])
      2 plt.colorbar()

NameError: name 'dim' is not defined

[33]:

segmentation.calculate_bg_iteratively(dim[0, 0].compute(), probplot=1)

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[33], line 1
----> 1 segmentation.calculate_bg_iteratively(dim[0, 0].compute(), probplot=1)

NameError: name 'dim' is not defined

[34]:

bgd[channels[0]][1].bg

---------------------------------------------------------------------------
KeyError                                  Traceback (most recent call last)
Cell In[34], line 1
----> 1 bgd[channels[0]][1].bg

KeyError: 'G'

[35]:

t = 0

plt.imshow(dim[t, 0] - bgd[channels[0]][t].bg)
plt.colorbar()

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[35], line 3
      1 t = 0
----> 3 plt.imshow(dim[t, 0] - bgd[channels[0]][t].bg)
      4 plt.colorbar()

NameError: name 'dim' is not defined

[36]:

m = segmentation.prob(dim[3, 0].compute(), 0, bgd[channels[0]][3].sd * 13) > 0.001
plt.imshow(~m)

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[36], line 1
----> 1 m = segmentation.prob(dim[3, 0].compute(), 0, bgd[channels[0]][3].sd * 13) > 0.001
      2 plt.imshow(~m)

NameError: name 'dim' is not defined

[37]:

asizeof(dim) / 1024**2

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[37], line 1
----> 1 asizeof(dim) / 1024**2

NameError: name 'dim' is not defined

[38]:

fpm = "/home/dan/workspace/nima_io/tests/data/t4_1.tif"
tifffile.TiffFile(fpm).series

---------------------------------------------------------------------------
FileNotFoundError                         Traceback (most recent call last)
/tmp/ipykernel_2661/1807629613.py in ?()
      1 fpm = "/home/dan/workspace/nima_io/tests/data/t4_1.tif"
----> 2 tifffile.TiffFile(fpm).series

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size, omexml, _multifile, _useframes, _parent, **is_flags)
   4240                 raise ValueError('invalid OME-XML')
   4241             self._omexml = omexml
   4242             self.is_ome = True
   4243
-> 4244         fh = FileHandle(file, mode=mode, name=name, offset=offset, size=size)
   4245         self._fh = fh
   4246         self._multifile = True if _multifile is None else bool(_multifile)
   4247         self._files = {fh.name: self}

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size)
  13350         self._offset = -1 if offset is None else offset
  13351         self._size = -1 if size is None else size
  13352         self._close = True
  13353         self._lock = NullContext()
> 13354         self.open()
  13355         assert self._fh is not None

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self)
  13369             if self._mode not in {'rb', 'r+b', 'wb', 'xb'}:
  13370                 raise ValueError(f'invalid mode {self._mode}')
  13371             self._file = os.path.realpath(self._file)
  13372             self._dir, self._name = os.path.split(self._file)
> 13373             self._fh = open(self._file, self._mode, encoding=None)
  13374             self._close = True
  13375             self._offset = max(0, self._offset)
  13376         elif isinstance(self._file, FileHandle):

FileNotFoundError: [Errno 2] No such file or directory: '/home/dan/workspace/nima_io/tests/data/t4_1.tif'

[39]:

md = nima.Metadata(tifffile.TiffReader(fpm))
md

---------------------------------------------------------------------------
FileNotFoundError                         Traceback (most recent call last)
/tmp/ipykernel_2661/1703928088.py in ?()
----> 1 md = nima.Metadata(tifffile.TiffReader(fpm))
      2 md

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size, omexml, _multifile, _useframes, _parent, **is_flags)
   4240                 raise ValueError('invalid OME-XML')
   4241             self._omexml = omexml
   4242             self.is_ome = True
   4243
-> 4244         fh = FileHandle(file, mode=mode, name=name, offset=offset, size=size)
   4245         self._fh = fh
   4246         self._multifile = True if _multifile is None else bool(_multifile)
   4247         self._files = {fh.name: self}

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size)
  13350         self._offset = -1 if offset is None else offset
  13351         self._size = -1 if size is None else size
  13352         self._close = True
  13353         self._lock = NullContext()
> 13354         self.open()
  13355         assert self._fh is not None

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self)
  13369             if self._mode not in {'rb', 'r+b', 'wb', 'xb'}:
  13370                 raise ValueError(f'invalid mode {self._mode}')
  13371             self._file = os.path.realpath(self._file)
  13372             self._dir, self._name = os.path.split(self._file)
> 13373             self._fh = open(self._file, self._mode, encoding=None)
  13374             self._close = True
  13375             self._offset = max(0, self._offset)
  13376         elif isinstance(self._file, FileHandle):

FileNotFoundError: [Errno 2] No such file or directory: '/home/dan/workspace/nima_io/tests/data/t4_1.tif'

[40]:

nima.read_tiffmd(fpm, ["G", "R2", "C", "R"])

---------------------------------------------------------------------------
FileNotFoundError                         Traceback (most recent call last)
/tmp/ipykernel_2661/143293133.py in ?()
----> 1 nima.read_tiffmd(fpm, ["G", "R2", "C", "R"])

~/work/nima/nima/src/nima/nima.py in ?(fp, channels)
   1072 def read_tiffmd(fp: Path, channels: Sequence[str]) -> tuple[Array, Metadata]:
   1073     """Read multichannel TIFF timelapse image."""
   1074     n_channels = len(channels)
-> 1075     rdr = TiffReader(fp)
   1076     dim = da.from_zarr(rdr.aszarr())  # type: ignore[no-untyped-call]
   1077     md = Metadata(rdr)
   1078     if md.size_c[0] % n_channels:

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size, omexml, _multifile, _useframes, _parent, **is_flags)
   4240                 raise ValueError('invalid OME-XML')
   4241             self._omexml = omexml
   4242             self.is_ome = True
   4243
-> 4244         fh = FileHandle(file, mode=mode, name=name, offset=offset, size=size)
   4245         self._fh = fh
   4246         self._multifile = True if _multifile is None else bool(_multifile)
   4247         self._files = {fh.name: self}

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size)
  13350         self._offset = -1 if offset is None else offset
  13351         self._size = -1 if size is None else size
  13352         self._close = True
  13353         self._lock = NullContext()
> 13354         self.open()
  13355         assert self._fh is not None

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self)
  13369             if self._mode not in {'rb', 'r+b', 'wb', 'xb'}:
  13370                 raise ValueError(f'invalid mode {self._mode}')
  13371             self._file = os.path.realpath(self._file)
  13372             self._dir, self._name = os.path.split(self._file)
> 13373             self._fh = open(self._file, self._mode, encoding=None)
  13374             self._close = True
  13375             self._offset = max(0, self._offset)
  13376         elif isinstance(self._file, FileHandle):

FileNotFoundError: [Errno 2] No such file or directory: '/home/dan/workspace/nima_io/tests/data/t4_1.tif'

[41]:

asizeof(dim) / 1024**2

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[41], line 1
----> 1 asizeof(dim) / 1024**2

NameError: name 'dim' is not defined

[42]:

bgr = segmentation.calculate_bg_iteratively(dim[1, 0].compute())
bgr.bg

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[42], line 1
----> 1 bgr = segmentation.calculate_bg_iteratively(dim[1, 0].compute())
      2 bgr.bg

NameError: name 'dim' is not defined

[ ]:

[43]:

dim, md = nima.read_tiffmd(fp_mdtest, channels=["G", "R2", "C", "R"])

md

---------------------------------------------------------------------------
FileNotFoundError                         Traceback (most recent call last)
/tmp/ipykernel_2661/2107487587.py in ?()
----> 1 dim, md = nima.read_tiffmd(fp_mdtest, channels=["G", "R2", "C", "R"])
      2
      3 md

~/work/nima/nima/src/nima/nima.py in ?(fp, channels)
   1072 def read_tiffmd(fp: Path, channels: Sequence[str]) -> tuple[Array, Metadata]:
   1073     """Read multichannel TIFF timelapse image."""
   1074     n_channels = len(channels)
-> 1075     rdr = TiffReader(fp)
   1076     dim = da.from_zarr(rdr.aszarr())  # type: ignore[no-untyped-call]
   1077     md = Metadata(rdr)
   1078     if md.size_c[0] % n_channels:

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size, omexml, _multifile, _useframes, _parent, **is_flags)
   4240                 raise ValueError('invalid OME-XML')
   4241             self._omexml = omexml
   4242             self.is_ome = True
   4243
-> 4244         fh = FileHandle(file, mode=mode, name=name, offset=offset, size=size)
   4245         self._fh = fh
   4246         self._multifile = True if _multifile is None else bool(_multifile)
   4247         self._files = {fh.name: self}

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size)
  13350         self._offset = -1 if offset is None else offset
  13351         self._size = -1 if size is None else size
  13352         self._close = True
  13353         self._lock = NullContext()
> 13354         self.open()
  13355         assert self._fh is not None

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self)
  13369             if self._mode not in {'rb', 'r+b', 'wb', 'xb'}:
  13370                 raise ValueError(f'invalid mode {self._mode}')
  13371             self._file = os.path.realpath(self._file)
  13372             self._dir, self._name = os.path.split(self._file)
> 13373             self._fh = open(self._file, self._mode, encoding=None)
  13374             self._close = True
  13375             self._offset = max(0, self._offset)
  13376         elif isinstance(self._file, FileHandle):

FileNotFoundError: [Errno 2] No such file or directory: '/home/dati/dt-clop3/data/ionofori assay pH6.8 22.05.05/Clop FLATxy Gexp20ms Rexp3ms.tf8'

[44]:

channels3 = ["G", "R", "C", "G2"]
dim3, _, _ = nima.read_tiff(fp_mdtest, channels3)

---------------------------------------------------------------------------
FileNotFoundError                         Traceback (most recent call last)
/tmp/ipykernel_2661/3014772201.py in ?()
      1 channels3 = ["G", "R", "C", "G2"]
----> 2 dim3, _, _ = nima.read_tiff(fp_mdtest, channels3)

~/work/nima/nima/src/nima/nima.py in ?(fp, channels)
     70     (3, 4)
     71
     72     """
     73     n_channels = len(channels)
---> 74     with TiffFile(fp) as tif:
     75         im = tif.asarray()
     76         axes = tif.series[0].axes
     77     idx = axes.rfind("T")

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size, omexml, _multifile, _useframes, _parent, **is_flags)
   4240                 raise ValueError('invalid OME-XML')
   4241             self._omexml = omexml
   4242             self.is_ome = True
   4243
-> 4244         fh = FileHandle(file, mode=mode, name=name, offset=offset, size=size)
   4245         self._fh = fh
   4246         self._multifile = True if _multifile is None else bool(_multifile)
   4247         self._files = {fh.name: self}

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size)
  13350         self._offset = -1 if offset is None else offset
  13351         self._size = -1 if size is None else size
  13352         self._close = True
  13353         self._lock = NullContext()
> 13354         self.open()
  13355         assert self._fh is not None

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self)
  13369             if self._mode not in {'rb', 'r+b', 'wb', 'xb'}:
  13370                 raise ValueError(f'invalid mode {self._mode}')
  13371             self._file = os.path.realpath(self._file)
  13372             self._dir, self._name = os.path.split(self._file)
> 13373             self._fh = open(self._file, self._mode, encoding=None)
  13374             self._close = True
  13375             self._offset = max(0, self._offset)
  13376         elif isinstance(self._file, FileHandle):

FileNotFoundError: [Errno 2] No such file or directory: '/home/dati/dt-clop3/data/ionofori assay pH6.8 22.05.05/Clop FLATxy Gexp20ms Rexp3ms.tf8'

[45]:

dim3, bgv3, bgf3 = nima.d_bg(
    dim3, segmentation.BgParams(kind="li_adaptive"), downscale=(2, 2)
)
c = dim3["C"]
g = dim3["G"]
r = dim3["R"]

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[45], line 2
      1 dim3, bgv3, bgf3 = nima.d_bg(
----> 2     dim3, segmentation.BgParams(kind="li_adaptive"), downscale=(2, 2)
      3 )
      4 c = dim3["C"]
      5 g = dim3["G"]

NameError: name 'dim3' is not defined

[46]:

bc = bgv3["C"]
bg = bgv3["G"][3]
br = bgv3["R"][3]

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[46], line 1
----> 1 bc = bgv3["C"]
      2 bg = bgv3["G"][3]
      3 br = bgv3["R"][3]

NameError: name 'bgv3' is not defined

[47]:

skimage.io.imshow(dim3["R"][1])

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[47], line 1
----> 1 skimage.io.imshow(dim3["R"][1])

NameError: name 'dim3' is not defined

[48]:

plt.plot(dim3["R"][1][80, 30:])

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[48], line 1
----> 1 plt.plot(dim3["R"][1][80, 30:])

NameError: name 'dim3' is not defined

[49]:

green = dim[0].compute()
bgg = segmentation.calculate_bg_iteratively(green)
bgg

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[49], line 1
----> 1 green = dim[0].compute()
      2 bgg = segmentation.calculate_bg_iteratively(green)
      3 bgg

NameError: name 'dim' is not defined

[50]:

mask = segmentation.prob(green, bgg.bg, bgg.sd) > 0.01
green[mask]

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[50], line 1
----> 1 mask = segmentation.prob(green, bgg.bg, bgg.sd) > 0.01
      2 green[mask]

NameError: name 'green' is not defined

[51]:

cyan = dim[2].compute()
bgc = segmentation.calculate_bg_iteratively(cyan)
mask = segmentation.prob(cyan, bgc.bg, bgc.sd) < 0.00000000000000000000001

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[51], line 1
----> 1 cyan = dim[2].compute()
      2 bgc = segmentation.calculate_bg_iteratively(cyan)
      3 mask = segmentation.prob(cyan, bgc.bg, bgc.sd) < 0.00000000000000000000001

NameError: name 'dim' is not defined

[52]:

red = dim[3].compute()
red2 = dim[1].compute()

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[52], line 1
----> 1 red = dim[3].compute()
      2 red2 = dim[1].compute()

NameError: name 'dim' is not defined

[53]:

plt.figure(figsize=(7.5, 7.5))

plt.subplot(3, 2, 1)
plt.hexbin(green[mask], cyan[mask], bins="log", cmap=plt.cm.viridis_r)
cb = plt.colorbar()
plt.xlabel("green")
plt.ylabel("cyan")

plt.subplot(3, 2, 2)
plt.hexbin(red[mask], cyan[mask], bins="log", cmap=plt.cm.viridis_r)
cb = plt.colorbar()
plt.xlabel("red")
plt.ylabel("cyan")

ax = plt.subplot(3, 2, 4)
plt.hexbin(red[mask], green[mask], bins="log", cmap=plt.cm.viridis_r)
cb = plt.colorbar()
plt.xlabel("red")
plt.ylabel("green")

ax = plt.subplot(3, 2, 5)
plt.hexbin(red2[mask], green[mask], bins="log", cmap=plt.cm.viridis_r)
cb = plt.colorbar()
plt.xlabel("red2")
plt.ylabel("green")

ax = plt.subplot(3, 2, 6)
plt.hexbin(red2[mask], cyan[mask], bins="log", cmap=plt.cm.viridis_r)
cb = plt.colorbar()
plt.xlabel("red2")
plt.ylabel("cyan")

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[53], line 4
      1 plt.figure(figsize=(7.5, 7.5))
      3 plt.subplot(3, 2, 1)
----> 4 plt.hexbin(green[mask], cyan[mask], bins="log", cmap=plt.cm.viridis_r)
      5 cb = plt.colorbar()
      6 plt.xlabel("green")

NameError: name 'green' is not defined

../_images/tutorials_usage_57_1.png

[54]:

from mpl_toolkits.axes_grid1.inset_locator import mark_inset

plt.figure(figsize=(10, 8))
plt.subplot(2, 2, 1)
plt.hexbin(g.ravel(), c.ravel(), bins="log", cmap=plt.cm.viridis_r)
cb = plt.colorbar()
plt.xlabel("green")
plt.ylabel("cyan")

plt.subplot(2, 2, 2)
plt.hexbin(r.ravel(), c.ravel(), bins="log", cmap=plt.cm.viridis_r)
cb = plt.colorbar()
plt.xlabel("red")
plt.ylabel("cyan")

ax = plt.subplot(2, 2, 4)
plt.hexbin(r.ravel(), g.ravel(), bins="log", cmap=plt.cm.viridis_r)
cb = plt.colorbar()
plt.xlabel("red")
plt.ylabel("green")

axins = plt.axes([0.2, 0.12, 0.28, 0.28])
axins.hexbin(
    r.ravel(), g.ravel(), extent=(0, 80, 0, 150), bins="log", cmap=plt.cm.viridis_r
)

mark_inset(ax, axins, loc1=1, loc2=4, fc="none", ec="0.5")

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[54], line 5
      3 plt.figure(figsize=(10, 8))
      4 plt.subplot(2, 2, 1)
----> 5 plt.hexbin(g.ravel(), c.ravel(), bins="log", cmap=plt.cm.viridis_r)
      6 cb = plt.colorbar()
      7 plt.xlabel("green")

NameError: name 'g' is not defined

../_images/tutorials_usage_58_1.png

[55]:

import statsmodels.api as sm

f = sm.qqplot_2samples(red[~mask], green[~mask])
f = sm.qqplot_2samples(red[~mask], cyan[~mask])

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[55], line 3
      1 import statsmodels.api as sm
----> 3 f = sm.qqplot_2samples(red[~mask], green[~mask])
      4 f = sm.qqplot_2samples(red[~mask], cyan[~mask])

NameError: name 'red' is not defined

1.3. flat image correction#

[56]:

g1a = nima.read_tiff("/home/dati/dt-clop3/data/210917/3alpha/c1_2.tf8", channels)[0][
    "G"
][2]
g2a = nima.read_tiff("/home/dati/dt-clop3/data/210917/3alpha/c2_2.tf8", channels)[0][
    "G"
][2]
g3a = nima.read_tiff("/home/dati/dt-clop3/data/210917/3alpha/c3_2.tf8", channels)[0][
    "G"
][2]
g1 = nima.read_tiff("/home/dati/dt-clop3/data/210917/3/c1_2.tf8", channels)[0]["G"][2]
g2 = nima.read_tiff("/home/dati/dt-clop3/data/210917/3/c2_2.tf8", channels)[0]["G"][2]
g3 = nima.read_tiff("/home/dati/dt-clop3/data/210917/3/c3_2.tf8", channels)[0]["G"][2]

---------------------------------------------------------------------------
FileNotFoundError                         Traceback (most recent call last)
/tmp/ipykernel_2661/4145832345.py in ?()
----> 1 g1a = nima.read_tiff("/home/dati/dt-clop3/data/210917/3alpha/c1_2.tf8", channels)[0][
      2     "G"
      3 ][2]
      4 g2a = nima.read_tiff("/home/dati/dt-clop3/data/210917/3alpha/c2_2.tf8", channels)[0][

~/work/nima/nima/src/nima/nima.py in ?(fp, channels)
     70     (3, 4)
     71
     72     """
     73     n_channels = len(channels)
---> 74     with TiffFile(fp) as tif:
     75         im = tif.asarray()
     76         axes = tif.series[0].axes
     77     idx = axes.rfind("T")

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size, omexml, _multifile, _useframes, _parent, **is_flags)
   4240                 raise ValueError('invalid OME-XML')
   4241             self._omexml = omexml
   4242             self.is_ome = True
   4243
-> 4244         fh = FileHandle(file, mode=mode, name=name, offset=offset, size=size)
   4245         self._fh = fh
   4246         self._multifile = True if _multifile is None else bool(_multifile)
   4247         self._files = {fh.name: self}

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size)
  13350         self._offset = -1 if offset is None else offset
  13351         self._size = -1 if size is None else size
  13352         self._close = True
  13353         self._lock = NullContext()
> 13354         self.open()
  13355         assert self._fh is not None

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self)
  13369             if self._mode not in {'rb', 'r+b', 'wb', 'xb'}:
  13370                 raise ValueError(f'invalid mode {self._mode}')
  13371             self._file = os.path.realpath(self._file)
  13372             self._dir, self._name = os.path.split(self._file)
> 13373             self._fh = open(self._file, self._mode, encoding=None)
  13374             self._close = True
  13375             self._offset = max(0, self._offset)
  13376         elif isinstance(self._file, FileHandle):

FileNotFoundError: [Errno 2] No such file or directory: '/home/dati/dt-clop3/data/210917/3alpha/c1_2.tf8'

[57]:

plt.subplot(1, 2, 1)
skimage.io.imshow(
    skimage.filters.gaussian(np.max([g1, g2, g3, g1a, g2a, g3a], axis=0), sigma=500)
)
plt.subplot(1, 2, 2)
skimage.io.imshow(skimage.filters.gaussian(np.max([g1, g2, g3], axis=0), sigma=500))

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[57], line 3
      1 plt.subplot(1, 2, 1)
      2 skimage.io.imshow(
----> 3     skimage.filters.gaussian(np.max([g1, g2, g3, g1a, g2a, g3a], axis=0), sigma=500)
      4 )
      5 plt.subplot(1, 2, 2)
      6 skimage.io.imshow(skimage.filters.gaussian(np.max([g1, g2, g3], axis=0), sigma=500))

NameError: name 'g1' is not defined

../_images/tutorials_usage_62_1.png

1.4. example w/out @Gain#

[58]:

fp24 = "/home/dati/dt-clop3/data/210917/3/c1_2.tf8"

[59]:

dim, md = nima.read_tiffmd(fp24, channels=["G", "R", "C"])

---------------------------------------------------------------------------
FileNotFoundError                         Traceback (most recent call last)
/tmp/ipykernel_2661/2668091231.py in ?()
----> 1 dim, md = nima.read_tiffmd(fp24, channels=["G", "R", "C"])

~/work/nima/nima/src/nima/nima.py in ?(fp, channels)
   1072 def read_tiffmd(fp: Path, channels: Sequence[str]) -> tuple[Array, Metadata]:
   1073     """Read multichannel TIFF timelapse image."""
   1074     n_channels = len(channels)
-> 1075     rdr = TiffReader(fp)
   1076     dim = da.from_zarr(rdr.aszarr())  # type: ignore[no-untyped-call]
   1077     md = Metadata(rdr)
   1078     if md.size_c[0] % n_channels:

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size, omexml, _multifile, _useframes, _parent, **is_flags)
   4240                 raise ValueError('invalid OME-XML')
   4241             self._omexml = omexml
   4242             self.is_ome = True
   4243
-> 4244         fh = FileHandle(file, mode=mode, name=name, offset=offset, size=size)
   4245         self._fh = fh
   4246         self._multifile = True if _multifile is None else bool(_multifile)
   4247         self._files = {fh.name: self}

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self, file, mode, name, offset, size)
  13350         self._offset = -1 if offset is None else offset
  13351         self._size = -1 if size is None else size
  13352         self._close = True
  13353         self._lock = NullContext()
> 13354         self.open()
  13355         assert self._fh is not None

~/work/nima/nima/.venv/lib/python3.13/site-packages/tifffile/tifffile.py in ?(self)
  13369             if self._mode not in {'rb', 'r+b', 'wb', 'xb'}:
  13370                 raise ValueError(f'invalid mode {self._mode}')
  13371             self._file = os.path.realpath(self._file)
  13372             self._dir, self._name = os.path.split(self._file)
> 13373             self._fh = open(self._file, self._mode, encoding=None)
  13374             self._close = True
  13375             self._offset = max(0, self._offset)
  13376         elif isinstance(self._file, FileHandle):

FileNotFoundError: [Errno 2] No such file or directory: '/home/dati/dt-clop3/data/210917/3/c1_2.tf8'

[60]:

sns.histplot(dim[8, 0].ravel())
plt.yscale("log")

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[60], line 1
----> 1 sns.histplot(dim[8, 0].ravel())
      2 plt.yscale("log")

NameError: name 'dim' is not defined

[61]:

utils.bg(dim[8, 0].compute())

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[61], line 1
----> 1 utils.bg(dim[8, 0].compute())

NameError: name 'dim' is not defined

[62]:

bgr = segmentation.calculate_bg_iteratively(dim[8, 1].compute())
bgr

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[62], line 1
----> 1 bgr = segmentation.calculate_bg_iteratively(dim[8, 1].compute())
      2 bgr

NameError: name 'dim' is not defined

[63]:

sns.histplot(
    dim[8, 1].compute()[
        (segmentation.prob(dim[8, 1].compute(), bgr.bg, bgr.sd) > 0.001)
    ]
)

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[63], line 2
      1 sns.histplot(
----> 2     dim[8, 1].compute()[
      3         (segmentation.prob(dim[8, 1].compute(), bgr.bg, bgr.sd) > 0.001)
      4     ]
      5 )

NameError: name 'dim' is not defined

[64]:

mask = (
    segmentation.prob(dim[8, 1].compute(), bgr.bg, bgr.sd)
    > 0.0000000000000000000000000000000000000000000000000000000000000000001
)
plt.imshow(~mask)

---------------------------------------------------------------------------
NameError                                 Traceback (most recent call last)
Cell In[64], line 2
      1 mask = (
----> 2     segmentation.prob(dim[8, 1].compute(), bgr.bg, bgr.sd)
      3     > 0.0000000000000000000000000000000000000000000000000000000000000000001
      4 )
      5 plt.imshow(~mask)

NameError: name 'dim' is not defined

1.5. BIAS and FLAT#

TODO:

pytest
build a function callable from both the library and the CLI