def segment_images(inpDir, outDir, config_data):
""" Workflow for data with a spotty appearance
in each 2d frame such as fibrillarin and beta catenin.
Args:
inpDir : path to the input directory
outDir : path to the output directory
config_data : path to the configuration file
"""
logging.basicConfig(
format='%(asctime)s - %(name)-8s - %(levelname)-8s - %(message)s',
datefmt='%d-%b-%y %H:%M:%S')
logger = logging.getLogger("main")
logger.setLevel(logging.INFO)
inpDir_files = os.listdir(inpDir)
for i, f in enumerate(inpDir_files):
logger.info('Segmenting image : {}'.format(f))
# Load image
br = BioReader(os.path.join(inpDir, f))
image = br.read_image()
structure_channel = 0
struct_img0 = image[:, :, :, structure_channel, 0]
struct_img0 = struct_img0.transpose(2, 0, 1).astype(np.float32)
# main algorithm
intensity_scaling_param = config_data['intensity_scaling_param']
struct_img = intensity_normalization(
struct_img0, scaling_param=intensity_scaling_param)
gaussian_smoothing_sigma = config_data['gaussian_smoothing_sigma']
structure_img_smooth = image_smoothing_gaussian_3d(
struct_img, sigma=gaussian_smoothing_sigma)
s2_param = config_data['s2_param']
bw = dot_2d_slice_by_slice_wrapper(structure_img_smooth, s2_param)
minArea = config_data['minArea']
seg = remove_small_objects(bw > 0,
min_size=minArea,
connectivity=1,
in_place=False)
seg = seg > 0
out_img = seg.astype(np.uint8)
out_img[out_img > 0] = 255
# create output image
out_img = out_img.transpose(1, 2, 0)
out_img = out_img.reshape(
(out_img.shape[0], out_img.shape[1], out_img.shape[2], 1, 1))
# write image using BFIO
bw = BioWriter(os.path.join(outDir, f))
bw.num_x(out_img.shape[1])
bw.num_y(out_img.shape[0])
bw.num_z(out_img.shape[2])
bw.num_c(out_img.shape[3])
bw.num_t(out_img.shape[4])
bw.pixel_type(dtype='uint8')
bw.write_image(out_img)
bw.close_image()
def segment_images(inpDir, outDir, config_data):
""" Workflow for data with similar morphology
as sialyltransferase 1.
Args:
inpDir : path to the input directory
outDir : path to the output directory
config_data : path to the configuration file
"""
logging.basicConfig(format='%(asctime)s - %(name)-8s - %(levelname)-8s - %(message)s',
datefmt='%d-%b-%y %H:%M:%S')
logger = logging.getLogger("main")
logger.setLevel(logging.INFO)
inpDir_files = os.listdir(inpDir)
for i,f in enumerate(inpDir_files):
logger.info('Segmenting image : {}'.format(f))
# Load image
br = BioReader(os.path.join(inpDir,f))
image = br.read_image()
structure_channel = 0
struct_img0 = image[:,:,:,structure_channel,0]
struct_img0 = struct_img0.transpose(2,0,1).astype(np.float32)
# main algorithm
intensity_scaling_param = config_data['intensity_scaling_param']
struct_img = intensity_normalization(struct_img0, scaling_param=intensity_scaling_param)
gaussian_smoothing_sigma = config_data['gaussian_smoothing_sigma']
structure_img_smooth = image_smoothing_gaussian_3d(struct_img, sigma=gaussian_smoothing_sigma)
global_thresh_method = config_data['global_thresh_method']
object_minArea = config_data['object_minArea']
bw, object_for_debug = MO(structure_img_smooth, global_thresh_method=global_thresh_method, object_minArea=object_minArea, return_object=True)
thin_dist_preserve = config_data['thin_dist_preserve']
thin_dist = config_data['thin_dist']
bw_thin = topology_preserving_thinning(bw>0, thin_dist_preserve, thin_dist)
s3_param = config_data['s3_param']
bw_extra = dot_3d_wrapper(structure_img_smooth, s3_param)
bw_combine = np.logical_or(bw_extra>0, bw_thin)
minArea = config_data['minArea']
seg = remove_small_objects(bw_combine>0, min_size=minArea, connectivity=1, in_place=False)
seg = seg > 0
out_img=seg.astype(np.uint8)
out_img[out_img>0]=255
# create output image
out_img = out_img.transpose(1,2,0)
out_img = out_img.reshape((out_img.shape[0], out_img.shape[1], out_img.shape[2], 1, 1))
# write image using BFIO
bw = BioWriter(os.path.join(outDir,f), metadata=br.read_metadata())
bw.num_x(out_img.shape[1])
bw.num_y(out_img.shape[0])
bw.num_z(out_img.shape[2])
bw.num_c(out_img.shape[3])
bw.num_t(out_img.shape[4])
bw.pixel_type(dtype='uint8')
bw.write_image(out_img)
bw.close_image()
def nuc_seg(image, scale, sigma, f2, hol_min, hol_max, minA):
img = intensity_normalization(image, scale)
img_s = image_smoothing_gaussian_3d(img, sigma)
bw = filament_2d_wrapper(img_s, f2)
bw_2 = hole_filling(bw, hol_min, hol_max)
seg = remove_small_objects(bw_2 > 0,
min_size=minA,
connectivity=1,
in_place=False)
seg2, n = label(seg, return_num=True)
return (img_s, bw_2, seg2, n)
def pex_seg(image, img_seg):
slices = []
Vol = []
pex_seg_img = []
pex_seg_n = []
slices = ndi.find_objects(img_seg)
img = intensity_normalization(image, scale)
img = image_smoothing_gaussian_3d(img, sigma)
for i, c in enumerate(slices):
cell_V = img_seg[c]
vt, f, n, val = marching_cubes_lewiner(cell_V)
Tvol = 1 / 6 * det(vt[f])
vol = abs(sum(Tvol))
Vol.append(vol)
cell = image[c]
cell_s = img[c]
bw = dot_3d_wrapper(cell_s, s3)
mask = remove_small_objects(bw > 0,
min_size=2,
connectivity=1,
in_place=False)
seed = dilation(peak_local_max(cell,
labels=label(mask),
min_distance=2,
indices=False),
selem=ball(1))
ws_map = -1 * ndi.distance_transform_edt(bw)
seg = watershed(ws_map, label(seed), mask=mask, watershed_line=True)
regions = regionprops(seg)
n = len(regions)
n = max(0, n)
pex_seg_img.append(seg)
pex_seg_n.append(n)
df = pd.DataFrame({
"Slices": slices,
"Cell Vol": Vol,
"Pex count": pex_seg_n
})
return df, pex_seg_img
def Workflow_cardio_npm1_100x(
struct_img: np.ndarray,
rescale_ratio: float = -1,
output_type: str = "default",
output_path: Union[str, Path] = None,
fn: Union[str, Path] = None,
output_func=None,
):
"""
classic segmentation workflow wrapper for structure Cardio NPM1 100x
Parameter:
-----------
struct_img: np.ndarray
the 3D image to be segmented
rescale_ratio: float
an optional parameter to allow rescale the image before running the
segmentation functions, default is no rescaling
output_type: str
select how to handle output. Currently, four types are supported:
1. default: the result will be saved at output_path whose filename is
original name without extention + "_struct_segmentaiton.tiff"
2. array: the segmentation result will be simply returned as a numpy array
3. array_with_contour: segmentation result will be returned together with
the contour of the segmentation
4. customize: pass in an extra output_func to do a special save. All the
intermediate results, names of these results, the output_path, and the
original filename (without extension) will be passed in to output_func.
"""
##########################################################################
# PARAMETERS:
# note that these parameters are supposed to be fixed for the structure
# and work well accross different datasets
intensity_norm_param = [0.5, 2.5]
gaussian_smoothing_sigma = 1
gaussian_smoothing_truncate_range = 3.0
dot_2d_sigma = 2
# dot_2d_sigma_extra = 1
# dot_2d_cutoff = 0.025
minArea = 1
low_level_min_size = 1000
##########################################################################
out_img_list = []
out_name_list = []
###################
# PRE_PROCESSING
###################
# intenisty normalization (min/max)
struct_img = intensity_normalization(struct_img,
scaling_param=intensity_norm_param)
out_img_list.append(struct_img.copy())
out_name_list.append("im_norm")
# rescale if needed
if rescale_ratio > 0:
struct_img = zoom(struct_img, (1, rescale_ratio, rescale_ratio),
order=2)
struct_img = (struct_img - struct_img.min() +
1e-8) / (struct_img.max() - struct_img.min() + 1e-8)
gaussian_smoothing_truncate_range = (
gaussian_smoothing_truncate_range * rescale_ratio)
# smoothing with gaussian filter
structure_img_smooth = image_smoothing_gaussian_3d(
struct_img,
sigma=gaussian_smoothing_sigma,
truncate_range=gaussian_smoothing_truncate_range,
)
out_img_list.append(structure_img_smooth.copy())
out_name_list.append("im_smooth")
###################
# core algorithm
###################
# step 1: low level thresholding
# global_otsu = threshold_otsu(structure_img_smooth)
global_tri = threshold_triangle(structure_img_smooth)
global_median = np.percentile(structure_img_smooth, 50)
th_low_level = (global_tri + global_median) / 2
# print(global_median)
# print(global_tri)
# print(th_low_level)
# imsave('img_smooth.tiff', structure_img_smooth)
bw_low_level = structure_img_smooth > th_low_level
bw_low_level = remove_small_objects(bw_low_level,
min_size=low_level_min_size,
connectivity=1,
in_place=True)
bw_low_level = dilation(bw_low_level, selem=ball(2))
# step 2: high level thresholding
local_cutoff = 0.333 * threshold_otsu(structure_img_smooth)
bw_high_level = np.zeros_like(bw_low_level)
lab_low, num_obj = label(bw_low_level, return_num=True, connectivity=1)
for idx in range(num_obj):
single_obj = lab_low == (idx + 1)
local_otsu = threshold_otsu(structure_img_smooth[single_obj])
if local_otsu > local_cutoff:
bw_high_level[np.logical_and(
structure_img_smooth > 1.2 * local_otsu, single_obj)] = 1
# imsave('seg_coarse.tiff', bw_high_level.astype(np.uint8))
out_img_list.append(bw_high_level.copy())
out_name_list.append("bw_coarse")
response_bright = dot_slice_by_slice(structure_img_smooth,
log_sigma=dot_2d_sigma)
bw_extra = response_bright > 0.03 # dot_2d_cutoff
bw_extra[~bw_low_level] = 0
bw_final = np.logical_or(bw_extra, bw_high_level)
# bw_final[holes]=0
###################
# POST-PROCESSING
###################
seg = remove_small_objects(bw_final,
min_size=minArea,
connectivity=1,
in_place=True)
# output
seg = seg > 0
seg = seg.astype(np.uint8)
seg[seg > 0] = 255
out_img_list.append(seg.copy())
out_name_list.append("bw_final")
if output_type == "default":
# the default final output, simply save it to the output path
save_segmentation(seg, False, Path(output_path), fn)
elif output_type == "customize":
# the hook for passing in a customized output function
# use "out_img_list" and "out_name_list" in your hook to
# customize your output functions
output_func(out_img_list, out_name_list, Path(output_path), fn)
elif output_type == "array":
return seg
elif output_type == "array_with_contour":
return (seg, generate_segmentation_contour(seg))
else:
raise NotImplementedError("invalid output type: {output_type}")
def Workflow_ctnnb1(
struct_img: np.ndarray,
rescale_ratio: float = -1,
output_type: str = "default",
output_path: Union[str, Path] = None,
fn: Union[str, Path] = None,
output_func=None,
):
"""
classic segmentation workflow wrapper for structure CTNNB1
Parameter:
-----------
struct_img: np.ndarray
the 3D image to be segmented
rescale_ratio: float
an optional parameter to allow rescale the image before running the
segmentation functions, default is no rescaling
output_type: str
select how to handle output. Currently, four types are supported:
1. default: the result will be saved at output_path whose filename is
original name without extention + "_struct_segmentaiton.tiff"
2. array: the segmentation result will be simply returned as a numpy array
3. array_with_contour: segmentation result will be returned together with
the contour of the segmentation
4. customize: pass in an extra output_func to do a special save. All the
intermediate results, names of these results, the output_path, and the
original filename (without extension) will be passed in to output_func.
"""
##########################################################################
# PARAMETERS:
# note that these parameters are supposed to be fixed for the structure
# and work well accross different datasets
intensity_norm_param = [4, 27]
gaussian_smoothing_sigma = 1
gaussian_smoothing_truncate_range = 3.0
dot_2d_sigma = 1.5
dot_2d_cutoff = 0.01
minArea = 10
##########################################################################
out_img_list = []
out_name_list = []
###################
# PRE_PROCESSING
###################
# intenisty normalization (min/max)
struct_img = intensity_normalization(struct_img,
scaling_param=intensity_norm_param)
out_img_list.append(struct_img.copy())
out_name_list.append("im_norm")
# rescale if needed
if rescale_ratio > 0:
struct_img = zoom(struct_img, (1, rescale_ratio, rescale_ratio),
order=2)
struct_img = (struct_img - struct_img.min() +
1e-8) / (struct_img.max() - struct_img.min() + 1e-8)
gaussian_smoothing_truncate_range = (
gaussian_smoothing_truncate_range * rescale_ratio)
# smoothing
structure_img_smooth = image_smoothing_gaussian_3d(
struct_img,
sigma=gaussian_smoothing_sigma,
truncate_range=gaussian_smoothing_truncate_range,
)
out_img_list.append(structure_img_smooth.copy())
out_name_list.append("im_smooth")
###################
# core algorithm
###################
response = dot_slice_by_slice(structure_img_smooth, log_sigma=dot_2d_sigma)
bw = response > dot_2d_cutoff
###################
# POST-PROCESSING
###################
seg = remove_small_objects(bw,
min_size=minArea,
connectivity=1,
in_place=False)
# output
seg = seg > 0
seg = seg.astype(np.uint8)
seg[seg > 0] = 255
out_img_list.append(seg.copy())
out_name_list.append("bw_final")
if output_type == "default":
# the default final output, simply save it to the output path
save_segmentation(seg, False, Path(output_path), fn)
elif output_type == "customize":
# the hook for passing in a customized output function
# use "out_img_list" and "out_name_list" in your hook to
# customize your output functions
output_func(out_img_list, out_name_list, Path(output_path), fn)
elif output_type == "array":
return seg
elif output_type == "array_with_contour":
return (seg, generate_segmentation_contour(seg))
else:
raise NotImplementedError("invalid output type: {output_type}")
def segment_images(inpDir, outDir, config_data):
""" Workflow for data with shell like shapes
such as lamin B1 (interphase-specific)
Args:
inpDir : path to the input directory
outDir : path to the output directory
config_data : path to the configuration file
"""
logging.basicConfig(
format='%(asctime)s - %(name)-8s - %(levelname)-8s - %(message)s',
datefmt='%d-%b-%y %H:%M:%S')
logger = logging.getLogger("main")
logger.setLevel(logging.INFO)
inpDir_files = os.listdir(inpDir)
for i, f in enumerate(inpDir_files):
logger.info('Segmenting image : {}'.format(f))
# Load image
br = BioReader(os.path.join(inpDir, f))
image = br.read_image()
structure_channel = 0
struct_img0 = image[:, :, :, structure_channel, 0]
struct_img0 = struct_img0.transpose(2, 0, 1).astype(np.float32)
# main algorithm
intensity_scaling_param = config_data['intensity_scaling_param']
struct_img = intensity_normalization(
struct_img0, scaling_param=intensity_scaling_param)
gaussian_smoothing_sigma = config_data['gaussian_smoothing_sigma']
structure_img_smooth = image_smoothing_gaussian_3d(
struct_img, sigma=gaussian_smoothing_sigma)
middle_frame_method = config_data['middle_frame_method']
mid_z = get_middle_frame(structure_img_smooth,
method=middle_frame_method)
f2_param = config_data['f2_param']
bw_mid_z = filament_2d_wrapper(structure_img_smooth[mid_z, :, :],
f2_param)
hole_max = config_data['hole_max']
hole_min = config_data['hole_min']
bw_fill_mid_z = hole_filling(bw_mid_z, hole_min, hole_max)
seed = get_3dseed_from_mid_frame(
np.logical_xor(bw_fill_mid_z, bw_mid_z), struct_img.shape, mid_z,
hole_min)
bw_filled = watershed(
struct_img, seed.astype(int), watershed_line=True) > 0
seg = np.logical_xor(bw_filled, dilation(bw_filled, selem=ball(1)))
seg = seg > 0
out_img = seg.astype(np.uint8)
out_img[out_img > 0] = 255
# create output image
out_img = out_img.transpose(1, 2, 0)
out_img = out_img.reshape(
(out_img.shape[0], out_img.shape[1], out_img.shape[2], 1, 1))
# write image using BFIO
bw = BioWriter(os.path.join(outDir, f))
bw.num_x(out_img.shape[1])
bw.num_y(out_img.shape[0])
bw.num_z(out_img.shape[2])
bw.num_c(out_img.shape[3])
bw.num_t(out_img.shape[4])
bw.pixel_type(dtype='uint8')
bw.write_image(out_img)
bw.close_image()
def Workflow_lmnb1_mitotic(struct_img,
rescale_ratio,
output_type,
output_path,
fn,
output_func=None):
##########################################################################
# PARAMETERS:
# note that these parameters are supposed to be fixed for the structure
# and work well accross different datasets
intensity_norm_param = [4000]
gaussian_smoothing_sigma = 1
gaussian_smoothing_truncate_range = 3.0
f2_param = [[0.5, 0.01]]
minArea = 5
##########################################################################
out_img_list = []
out_name_list = []
###################
# PRE_PROCESSING
###################
# intenisty normalization (min/max)
struct_img = intensity_normalization(struct_img,
scaling_param=intensity_norm_param)
out_img_list.append(struct_img.copy())
out_name_list.append('im_norm')
# rescale if needed
if rescale_ratio > 0:
struct_img = resize(struct_img, [1, rescale_ratio, rescale_ratio],
method="cubic")
struct_img = (struct_img - struct_img.min() +
1e-8) / (struct_img.max() - struct_img.min() + 1e-8)
# smoothing with boundary preserving smoothing
structure_img_smooth = image_smoothing_gaussian_3d(
struct_img,
sigma=gaussian_smoothing_sigma,
truncate_range=gaussian_smoothing_truncate_range)
out_img_list.append(structure_img_smooth.copy())
out_name_list.append('im_smooth')
###################
# core algorithm
###################
# 2d filament filter on the middle frame
bw = filament_2d_wrapper(structure_img_smooth, f2_param)
###################
# POST-PROCESSING
###################
bw = remove_small_objects(bw > 0,
min_size=minArea,
connectivity=1,
in_place=False)
# output
seg = bw > 0
seg = seg.astype(np.uint8)
seg[seg > 0] = 255
out_img_list.append(seg.copy())
out_name_list.append('bw_final')
if output_type == 'default':
# the default final output
save_segmentation(seg, False, output_path, fn)
elif output_type == 'array':
return seg
elif output_type == 'array_with_contour':
return (seg, generate_segmentation_contour(seg))
else:
print('your can implement your output hook here, but not yet')
quit()
def Workflow_fbl_labelfree_4dn(
struct_img: np.ndarray,
rescale_ratio: float = -1,
output_type: str = "default",
output_path: Union[str, Path] = None,
fn: Union[str, Path] = None,
output_func=None,
):
"""
classic segmentation workflow wrapper for structure FBL Labelfree 4dn
Parameter:
-----------
struct_img: np.ndarray
the 3D image to be segmented
rescale_ratio: float
an optional parameter to allow rescale the image before running the
segmentation functions, default is no rescaling
output_type: str
select how to handle output. Currently, four types are supported:
1. default: the result will be saved at output_path whose filename is
original name without extention + "_struct_segmentaiton.tiff"
2. array: the segmentation result will be simply returned as a numpy array
3. array_with_contour: segmentation result will be returned together with
the contour of the segmentation
4. customize: pass in an extra output_func to do a special save. All the
intermediate results, names of these results, the output_path, and the
original filename (without extension) will be passed in to output_func.
"""
##########################################################################
# PARAMETERS:
minArea = 5
low_level_min_size = 7000
s2_param = [[0.5, 0.1]]
intensity_scaling_param = [0.5, 19.5]
gaussian_smoothing_sigma = 1
##########################################################################
out_img_list = []
out_name_list = []
###################
# PRE_PROCESSING
###################
# intenisty normalization
struct_norm = intensity_normalization(
struct_img, scaling_param=intensity_scaling_param)
out_img_list.append(struct_img.copy())
out_name_list.append("im_norm")
# smoothing
struct_smooth = image_smoothing_gaussian_3d(struct_norm,
sigma=gaussian_smoothing_sigma)
out_img_list.append(struct_smooth.copy())
out_name_list.append("im_smooth")
###################
# core algorithm
###################
# step 1: low level thresholding
# global_otsu = threshold_otsu(structure_img_smooth)
global_tri = threshold_triangle(struct_smooth)
global_median = np.percentile(struct_smooth, 50)
th_low_level = (global_tri + global_median) / 2
bw_low_level = struct_smooth > th_low_level
bw_low_level = remove_small_objects(bw_low_level,
min_size=low_level_min_size,
connectivity=1,
in_place=True)
# step 2: high level thresholding
bw_high_level = np.zeros_like(bw_low_level)
lab_low, num_obj = label(bw_low_level, return_num=True, connectivity=1)
for idx in range(num_obj):
single_obj = lab_low == (idx + 1)
local_otsu = threshold_otsu(struct_smooth[single_obj])
bw_high_level[np.logical_and(struct_smooth > local_otsu * 1.2,
single_obj)] = 1
# step 3: finer segmentation
response2d = dot_2d_slice_by_slice_wrapper(struct_smooth, s2_param)
bw_finer = remove_small_objects(response2d,
min_size=minArea,
connectivity=1,
in_place=True)
# merge finer level detection into high level coarse segmentation
# to include outside dim parts
bw_high_level[bw_finer > 0] = 1
###################
# POST-PROCESSING
# make sure the variable name of final segmentation is 'seg'
###################
seg = remove_small_objects(bw_high_level,
min_size=minArea,
connectivity=1,
in_place=True)
# output
seg = seg > 0
seg = seg.astype(np.uint8)
seg[seg > 0] = 255
out_img_list.append(seg.copy())
out_name_list.append("bw_final")
if output_type == "default":
# the default final output, simply save it to the output path
save_segmentation(seg, False, Path(output_path), fn)
elif output_type == "customize":
# the hook for passing in a customized output function
# use "out_img_list" and "out_name_list" in your hook to
# customize your output functions
# TODO CREATE OUT_IMG_LIST/OUT_NAMELIST?
output_func(out_img_list, out_name_list, Path(output_path), fn)
elif output_type == "array":
return seg
elif output_type == "array_with_contour":
return (seg, generate_segmentation_contour(seg))
else:
raise NotImplementedError("invalid output type: {output_type}")
# get voxel dimensions
dim_iter = mtree.find('Metadata').find('Scaling').find('Items').findall('Distance')
dims = {}
for dimension in dim_iter:
dim_id = dimension.get('Id')
dims.update({dim_id:float(dimension.find('Value').text)*1.E6}) # [um]
voxel_vol = dims['X'] * dims['Y'] * dims['Z'] # [um^3]
dims_xyz = np.array([dims['X'], dims['Y'], dims['Z']])
#-- SEGMENT SPOTS -----------------------------------------------------------#
for i, g in enumerate(genes):
print('\nProcessing gene ' + g + '...\n')
print('Smoothing image...')
img_rna_smooth = image_smoothing_gaussian_3d(imgs_rna[i], sigma=1)
s3_param = [[1, 0.75*t_spots[i]], [2, 0.75*t_spots[i]]]
print('Identifying spots...')
bw = dot_3d_wrapper(img_rna_smooth, s3_param)
# watershed
minArea = 4
Mask = morphology.remove_small_objects(bw>0, min_size=minArea, connectivity=1, in_place=False)
labeled_mask = measure.label(Mask)
print('Performing watershed segmentation...')
peaks = feature.peak_local_max(imgs_rna[i],labels=labeled_mask, min_distance=2, indices=False)
Seed = morphology.binary_dilation(peaks, selem=morphology.ball(1))
Watershed_Map = -1*distance_transform_edt(bw)
seg = morphology.watershed(Watershed_Map, measure.label(Seed), mask=Mask, watershed_line=True)
seg = morphology.remove_small_objects(seg>0, min_size=minArea, connectivity=1, in_place=False)
def Workflow_cardio_npm1_100x(struct_img,
rescale_ratio,
output_type,
output_path,
fn,
output_func=None):
##########################################################################
# PARAMETERS:
# note that these parameters are supposed to be fixed for the structure
# and work well accross different datasets
intensity_norm_param = [0.5, 2.5]
gaussian_smoothing_sigma = 1
gaussian_smoothing_truncate_range = 3.0
dot_2d_sigma = 2
dot_2d_sigma_extra = 1
dot_2d_cutoff = 0.025
minArea = 1
low_level_min_size = 1000
##########################################################################
out_img_list = []
out_name_list = []
###################
# PRE_PROCESSING
###################
# intenisty normalization (min/max)
struct_img = intensity_normalization(struct_img,
scaling_param=intensity_norm_param)
out_img_list.append(struct_img.copy())
out_name_list.append('im_norm')
# rescale if needed
if rescale_ratio > 0:
struct_img = resize(struct_img, [1, rescale_ratio, rescale_ratio],
method="cubic")
struct_img = (struct_img - struct_img.min() +
1e-8) / (struct_img.max() - struct_img.min() + 1e-8)
gaussian_smoothing_truncate_range = gaussian_smoothing_truncate_range * rescale_ratio
# smoothing with gaussian filter
structure_img_smooth = image_smoothing_gaussian_3d(
struct_img,
sigma=gaussian_smoothing_sigma,
truncate_range=gaussian_smoothing_truncate_range)
out_img_list.append(structure_img_smooth.copy())
out_name_list.append('im_smooth')
###################
# core algorithm
###################
# step 1: low level thresholding
#global_otsu = threshold_otsu(structure_img_smooth)
global_tri = threshold_triangle(structure_img_smooth)
global_median = np.percentile(structure_img_smooth, 50)
th_low_level = (global_tri + global_median) / 2
#print(global_median)
#print(global_tri)
#print(th_low_level)
#imsave('img_smooth.tiff', structure_img_smooth)
bw_low_level = structure_img_smooth > th_low_level
bw_low_level = remove_small_objects(bw_low_level,
min_size=low_level_min_size,
connectivity=1,
in_place=True)
bw_low_level = dilation(bw_low_level, selem=ball(2))
# step 2: high level thresholding
local_cutoff = 0.333 * threshold_otsu(structure_img_smooth)
bw_high_level = np.zeros_like(bw_low_level)
lab_low, num_obj = label(bw_low_level, return_num=True, connectivity=1)
for idx in range(num_obj):
single_obj = (lab_low == (idx + 1))
local_otsu = threshold_otsu(structure_img_smooth[single_obj])
if local_otsu > local_cutoff:
bw_high_level[np.logical_and(
structure_img_smooth > 1.2 * local_otsu, single_obj)] = 1
#imsave('seg_coarse.tiff', bw_high_level.astype(np.uint8))
out_img_list.append(bw_high_level.copy())
out_name_list.append('bw_coarse')
response_bright = dot_slice_by_slice(structure_img_smooth,
log_sigma=dot_2d_sigma)
'''
response_dark = dot_slice_by_slice(1 - structure_img_smooth, log_sigma=dot_2d_sigma)
response_dark_extra = dot_slice_by_slice(1 - structure_img_smooth, log_sigma=dot_2d_sigma_extra)
imsave('res_dark_1.tiff', response_dark)
imsave('res_dark_2.tiff', response_dark_extra)
#inner_mask = bw_high_level.copy()
#for zz in range(inner_mask.shape[0]):
# inner_mask[zz,:,:] = binary_fill_holes(inner_mask[zz,:,:])
holes = np.logical_or(response_dark>dot_2d_cutoff , response_dark_extra>dot_2d_cutoff)
#holes[~inner_mask] = 0
'''
bw_extra = response_bright > 0.03 # dot_2d_cutoff
bw_extra[~bw_low_level] = 0
bw_final = np.logical_or(bw_extra, bw_high_level)
#bw_final[holes]=0
###################
# POST-PROCESSING
###################
seg = remove_small_objects(bw_final,
min_size=minArea,
connectivity=1,
in_place=True)
# output
seg = seg > 0
seg = seg.astype(np.uint8)
seg[seg > 0] = 255
out_img_list.append(seg.copy())
out_name_list.append('bw_final')
if output_type == 'default':
# the default final output
save_segmentation(seg, False, output_path, fn)
elif output_type == 'AICS_pipeline':
# pre-defined output function for pipeline data
save_segmentation(seg, True, output_path, fn)
elif output_type == 'customize':
# the hook for passing in a customized output function
output_fun(out_img_list, out_name_list, output_path, fn)
elif output_type == 'array':
return seg
elif output_type == 'array_with_contour':
return (seg, generate_segmentation_contour(seg))
else:
# the hook for pre-defined RnD output functions (AICS internal)
img_list, name_list = NPM1_output(out_img_list, out_name_list,
output_type, output_path, fn)
if output_type == 'QCB':
return img_list, name_list
def segment_images(inpDir, outDir, config_data):
""" Workflow for dot like shapes such as
Centrin-2, Desmoplakin, PMP34.
Args:
inpDir : path to the input directory
outDir : path to the output directory
config_data : path to the configuration file
"""
logging.basicConfig(
format='%(asctime)s - %(name)-8s - %(levelname)-8s - %(message)s',
datefmt='%d-%b-%y %H:%M:%S')
logger = logging.getLogger("main")
logger.setLevel(logging.INFO)
inpDir_files = os.listdir(inpDir)
for i, f in enumerate(inpDir_files):
logger.info('Segmenting image : {}'.format(f))
# Load an image
br = BioReader(os.path.join(inpDir, f))
image = br.read_image()
structure_channel = 0
struct_img0 = image[:, :, :, structure_channel, 0]
struct_img0 = struct_img0.transpose(2, 0, 1).astype(np.float32)
# main algorithm
intensity_scaling_param = config_data['intensity_scaling_param']
struct_img = intensity_normalization(
struct_img0, scaling_param=intensity_scaling_param)
gaussian_smoothing_sigma = config_data['gaussian_smoothing_sigma']
if config_data["gaussian_smoothing"] == "gaussian_slice_by_slice":
structure_img_smooth = image_smoothing_gaussian_slice_by_slice(
struct_img, sigma=gaussian_smoothing_sigma)
else:
structure_img_smooth = image_smoothing_gaussian_3d(
struct_img, sigma=gaussian_smoothing_sigma)
s3_param = config_data['s3_param']
bw = dot_3d_wrapper(structure_img_smooth, s3_param)
minArea = config_data['minArea']
Mask = remove_small_objects(bw > 0,
min_size=minArea,
connectivity=1,
in_place=False)
Seed = dilation(peak_local_max(struct_img,
labels=label(Mask),
min_distance=2,
indices=False),
selem=ball(1))
Watershed_Map = -1 * distance_transform_edt(bw)
seg = watershed(Watershed_Map,
label(Seed),
mask=Mask,
watershed_line=True)
seg = remove_small_objects(seg > 0,
min_size=minArea,
connectivity=1,
in_place=False)
seg = seg > 0
out_img = seg.astype(np.uint8)
out_img[out_img > 0] = 255
# create output image
out_img = out_img.transpose(1, 2, 0)
out_img = out_img.reshape(
(out_img.shape[0], out_img.shape[1], out_img.shape[2], 1, 1))
# write image using BFIO
bw = BioWriter(os.path.join(outDir, f))
bw.num_x(out_img.shape[1])
bw.num_y(out_img.shape[0])
bw.num_z(out_img.shape[2])
bw.num_c(out_img.shape[3])
bw.num_t(out_img.shape[4])
bw.pixel_type(dtype='uint8')
bw.write_image(out_img)
bw.close_image()
def Workflow_lmnb1_interphase(struct_img,
rescale_ratio,
output_type,
output_path,
fn,
output_func=None):
##########################################################################
# PARAMETERS:
# note that these parameters are supposed to be fixed for the structure
# and work well accross different datasets
intensity_norm_param = [4000]
gaussian_smoothing_sigma = 1
gaussian_smoothing_truncate_range = 3.0
f2_param = [[1, 0.01], [2, 0.01], [3, 0.01]]
hole_max = 40000
hole_min = 400
##########################################################################
out_img_list = []
out_name_list = []
###################
# PRE_PROCESSING
###################
# intenisty normalization (min/max)
struct_img = intensity_normalization(struct_img,
scaling_param=intensity_norm_param)
out_img_list.append(struct_img.copy())
out_name_list.append('im_norm')
# rescale if needed
if rescale_ratio > 0:
struct_img = resize(struct_img, [1, rescale_ratio, rescale_ratio],
method="cubic")
struct_img = (struct_img - struct_img.min() +
1e-8) / (struct_img.max() - struct_img.min() + 1e-8)
# smoothing with boundary preserving smoothing
structure_img_smooth = image_smoothing_gaussian_3d(
struct_img,
sigma=gaussian_smoothing_sigma,
truncate_range=gaussian_smoothing_truncate_range)
out_img_list.append(structure_img_smooth.copy())
out_name_list.append('im_smooth')
###################
# core algorithm
###################
# get the mid frame
mid_z = get_middle_frame(structure_img_smooth, method='intensity')
# 2d filament filter on the middle frame
bw_mid_z = filament_2d_wrapper(structure_img_smooth[mid_z, :, :], f2_param)
# hole filling
bw_fill_mid_z = hole_filling(bw_mid_z, hole_min, hole_max)
seed = get_3dseed_from_mid_frame(np.logical_xor(bw_fill_mid_z, bw_mid_z),
struct_img.shape,
mid_z,
hole_min,
bg_seed=True)
seg_filled = watershed(
struct_img, seed.astype(int),
watershed_line=True) > 1 # in watershed result, 1 is for background
# get the actual shell
seg = find_boundaries(seg_filled, connectivity=1, mode='thick')
# output
seg = seg.astype(np.uint8)
seg[seg > 0] = 255
out_img_list.append(seg.copy())
out_name_list.append('bw_final')
if output_type == 'default':
# the default final output
save_segmentation(seg, False, output_path, fn)
elif output_type == 'array':
return seg
elif output_type == 'array_with_contour':
return (seg, generate_segmentation_contour(seg))
else:
print('your can implement your output hook here, but not yet')
quit()
def cell_mask(image, scale, sigma):
img1 = intensity_normalization(image, scale)
img_s = image_smoothing_gaussian_3d(img1, sigma)
thr = threshold_otsu(img_s)
bw = img_s > thr * 0.8
return (img_s, bw)
def Workflow_lmnb1_interphase(
struct_img: np.ndarray,
rescale_ratio: float = -1,
output_type: str = "default",
output_path: Union[str, Path] = None,
fn: Union[str, Path] = None,
output_func=None,
):
"""
classic segmentation workflow wrapper for structure LMNB1 interphase
Parameter:
-----------
struct_img: np.ndarray
the 3D image to be segmented
rescale_ratio: float
an optional parameter to allow rescale the image before running the
segmentation functions, default is no rescaling
output_type: str
select how to handle output. Currently, four types are supported:
1. default: the result will be saved at output_path whose filename is
original name without extention + "_struct_segmentaiton.tiff"
2. array: the segmentation result will be simply returned as a numpy array
3. array_with_contour: segmentation result will be returned together with
the contour of the segmentation
4. customize: pass in an extra output_func to do a special save. All the
intermediate results, names of these results, the output_path, and the
original filename (without extension) will be passed in to output_func.
"""
##########################################################################
# PARAMETERS:
# note that these parameters are supposed to be fixed for the structure
# and work well accross different datasets
intensity_norm_param = [4000]
gaussian_smoothing_sigma = 1
gaussian_smoothing_truncate_range = 3.0
f2_param = [[1, 0.01], [2, 0.01], [3, 0.01]]
hole_max = 40000
hole_min = 400
##########################################################################
out_img_list = []
out_name_list = []
###################
# PRE_PROCESSING
###################
# intenisty normalization (min/max)
struct_img = intensity_normalization(struct_img,
scaling_param=intensity_norm_param)
out_img_list.append(struct_img.copy())
out_name_list.append("im_norm")
# rescale if needed
if rescale_ratio > 0:
struct_img = zoom(struct_img, (1, rescale_ratio, rescale_ratio),
order=2)
struct_img = (struct_img - struct_img.min() +
1e-8) / (struct_img.max() - struct_img.min() + 1e-8)
# smoothing with boundary preserving smoothing
structure_img_smooth = image_smoothing_gaussian_3d(
struct_img,
sigma=gaussian_smoothing_sigma,
truncate_range=gaussian_smoothing_truncate_range,
)
out_img_list.append(structure_img_smooth.copy())
out_name_list.append("im_smooth")
###################
# core algorithm
###################
# get the mid frame
mid_z = get_middle_frame(structure_img_smooth, method="intensity")
# 2d filament filter on the middle frame
bw_mid_z = filament_2d_wrapper(structure_img_smooth[mid_z, :, :], f2_param)
# hole filling
bw_fill_mid_z = hole_filling(bw_mid_z, hole_min, hole_max)
seed = get_3dseed_from_mid_frame(
np.logical_xor(bw_fill_mid_z, bw_mid_z),
struct_img.shape,
mid_z,
hole_min,
bg_seed=True,
)
seg_filled = (
watershed(struct_img, seed.astype(int), watershed_line=True) > 1
) # in watershed result, 1 is for background
# get the actual shell
seg = find_boundaries(seg_filled, connectivity=1, mode="thick")
# output
seg = seg.astype(np.uint8)
seg[seg > 0] = 255
out_img_list.append(seg.copy())
out_name_list.append("bw_final")
if output_type == "default":
# the default final output, simply save it to the output path
save_segmentation(seg, False, Path(output_path), fn)
elif output_type == "customize":
# the hook for passing in a customized output function
# use "out_img_list" and "out_name_list" in your hook to
# customize your output functions
output_func(out_img_list, out_name_list, Path(output_path), fn)
elif output_type == "array":
return seg
elif output_type == "array_with_contour":
return (seg, generate_segmentation_contour(seg))
else:
raise NotImplementedError("invalid output type: {output_type}")
def Workflow_st6gal1(
struct_img: np.ndarray,
rescale_ratio: float = -1,
output_type: str = "default",
output_path: Union[str, Path] = None,
fn: Union[str, Path] = None,
output_func=None,
):
"""
classic segmentation workflow wrapper for structure ST6GAL1
Parameter:
-----------
struct_img: np.ndarray
the 3D image to be segmented
rescale_ratio: float
an optional parameter to allow rescale the image before running the
segmentation functions, default is no rescaling
output_type: str
select how to handle output. Currently, four types are supported:
1. default: the result will be saved at output_path whose filename is
original name without extention + "_struct_segmentaiton.tiff"
2. array: the segmentation result will be simply returned as a numpy array
3. array_with_contour: segmentation result will be returned together with
the contour of the segmentation
4. customize: pass in an extra output_func to do a special save. All the
intermediate results, names of these results, the output_path, and the
original filename (without extension) will be passed in to output_func.
"""
##########################################################################
# PARAMETERS:
# note that these parameters are supposed to be fixed for the structure
# and work well accross different datasets
intensity_norm_param = [9, 19]
gaussian_smoothing_sigma = 1
gaussian_smoothing_truncate_range = 3.0
cell_wise_min_area = 1200
dot_3d_sigma = 1.6
dot_3d_cutoff = 0.02
minArea = 10
thin_dist = 1
thin_dist_preserve = 1.6
##########################################################################
out_img_list = []
out_name_list = []
###################
# PRE_PROCESSING
###################
# intenisty normalization (min/max)
struct_img = intensity_normalization(struct_img, scaling_param=intensity_norm_param)
out_img_list.append(struct_img.copy())
out_name_list.append("im_norm")
# rescale if needed
if rescale_ratio > 0:
struct_img = zoom(struct_img, (1, rescale_ratio, rescale_ratio), order=2)
struct_img = (struct_img - struct_img.min() + 1e-8) / (
struct_img.max() - struct_img.min() + 1e-8
)
gaussian_smoothing_truncate_range = (
gaussian_smoothing_truncate_range * rescale_ratio
)
# smoothing with gaussian filter
structure_img_smooth = image_smoothing_gaussian_3d(
struct_img,
sigma=gaussian_smoothing_sigma,
truncate_range=gaussian_smoothing_truncate_range,
)
out_img_list.append(structure_img_smooth.copy())
out_name_list.append("im_smooth")
###################
# core algorithm
###################
# cell-wise local adaptive thresholding
th_low_level = threshold_triangle(structure_img_smooth)
bw_low_level = structure_img_smooth > th_low_level
bw_low_level = remove_small_objects(
bw_low_level, min_size=cell_wise_min_area, connectivity=1, in_place=True
)
bw_low_level = dilation(bw_low_level, selem=ball(2))
bw_high_level = np.zeros_like(bw_low_level)
lab_low, num_obj = label(bw_low_level, return_num=True, connectivity=1)
for idx in range(num_obj):
single_obj = lab_low == (idx + 1)
local_otsu = threshold_otsu(structure_img_smooth[single_obj > 0])
bw_high_level[
np.logical_and(structure_img_smooth > local_otsu * 0.98, single_obj)
] = 1
# LOG 3d to capture spots
response = dot_3d(structure_img_smooth, log_sigma=dot_3d_sigma)
bw_extra = response > dot_3d_cutoff
# thinning
bw_high_level = topology_preserving_thinning(
bw_high_level, thin_dist_preserve, thin_dist
)
# combine the two parts
bw = np.logical_or(bw_high_level, bw_extra)
###################
# POST-PROCESSING
###################
seg = remove_small_objects(bw > 0, min_size=minArea, connectivity=1, in_place=False)
# output
seg = seg > 0
seg = seg.astype(np.uint8)
seg[seg > 0] = 255
out_img_list.append(seg.copy())
out_name_list.append("bw_final")
if output_type == "default":
# the default final output, simply save it to the output path
save_segmentation(seg, False, Path(output_path), fn)
elif output_type == "customize":
# the hook for passing in a customized output function
# use "out_img_list" and "out_name_list" in your hook to
# customize your output functions
output_func(out_img_list, out_name_list, Path(output_path), fn)
elif output_type == "array":
return seg
elif output_type == "array_with_contour":
return (seg, generate_segmentation_contour(seg))
else:
raise NotImplementedError("invalid output type: {output_type}")
