+
+import pandas as pd
+import numpy as np
+import libtiff
+import skimage.exposure
+import skimage.feature
+import skimage.filters
+import skimage.morphology
+
+def make_mask(zstack, nbins=32768):
+ thresh = skimage.filters.threshold_otsu(zstack, nbins=32768)
+ return zstack > thresh
+
+def build_blob_array(stack):
+ data = [[[False for i in range(len(stack[0][0]))] for j in range(len(stack[0]))] for k in range(len(stack))]
+ for im in range(len(stack))): #was stack
+ print "processing " + str(im)
+ yx = skimage.exposure.equalize_adapthist(stack[im], clip_limit=0.00025, nbins=32768)
+
+ zmx = np.max(yx)
+ zmn = np.min(yx)
+ yx = (yx - zmn) / (zmx - zmn)
+
+ blobs_log = skimage.feature.blob_log(yx, min_sigma=1, max_sigma=3, num_sigma=20, threshold=0.025)
+ print str(len(blobs_log)) + " blobs found"
+ if len(blobs_log) > 1:
+ blobs_log[:, 2] = blobs_log[:, 2] * math.sqrt(2)
+ for i in blobs_log:
+ data[im][int(i[0])][int(i[1])] = True
+ return data
+
+def make_watershed(mask):
+ distance = ndi.distance_transform_edt(mask)
+ local_max = skimage.feature.peak_local_max(distance, indices=False, footprint=np.ones((31, 31)), labels=mask)
+ markers = ndi.label(local_max)[0]
+ labels = skimage.morphology.watershed(-distance, markers, mask=mask)
+ return labels
+
+def make_gene_layers(gene_tif):
+ gene_z = gene_tif.get_tiff_array()
+ gene_s = np.arange(0, len(gene_z)+1, len(gene_z) / 6)
+ gene_r = zip(gene_s[:-1], (gene_s + 1)[1:])
+ gene_y = np.array(gene_z[gene_r[2][0]:gene_r[2][1]], dtype=np.uint16)
+ return gene_y
+
+def make_nissl_watershed(nissl_tif):
+ nissl_z = nissl_tif.get_tiff_array()
+ nissl_s = np.arange(0, len(nissl_z)+1, len(nissl_z) / 6)
+ nissl_r = zip(nissl_s[:-1], (nissl_s + 1)[1:])
+ nissl_y = np.array(nissl_z[nissl_r[1][0]:nissl_r[1][1]], dtype=np.uint16)
+ nissl_watershed = [[] for k in range(len(nissl_y))]
+
+ for i in range(len(nissl_y)):
+ mask = make_mask(nissl_y[i])
+ watershed = make_watershed(mask)
+ nissl_watershed[i] = watershed
+ return nissl_watershed
+
+def correct_labels(nissl_watershed):
+ nissl_watershed_corrected = [[[0 for i in range(len(nissl_watershed[0][0]))] for j in range(len(nissl_watershed[0]))] for k in range(len(nissl_watershed))]
+ nissl_watershed_corrected[0] = [i for i in nissl_watershed[0]]
+ nissl_watershed_corrected = np.asarray(nissl_watershed_corrected)
+
+ for k in range(1, len(nissl_watershed_corrected)):
+ cell_number_max = nissl_watershed_corrected[k-1].max() #changed to corrected to keep accurate count
+ cell_number_two_max = nissl_watershed[k].max()
+ and_map = np.logical_and(nissl_watershed[k-1], nissl_watershed[k])
+ and_map_pos = np.where(and_map)
+ cell_set = set(nissl_watershed[k-1][np.where(and_map)])
+ inter_layer_dict = dict()
+ for i in range(len(and_map_pos[0])):
+ inter_layer_dict[nissl_watershed[k][and_map_pos[0][i]][and_map_pos[1][i]]] = nissl_watershed[k-1][and_map_pos[0][i]][and_map_pos[1][i]]
+ counter = 0
+ inter_layer_dict[0] = 0
+ for i in range(1, cell_number_two_max + 1):
+ if not i in inter_layer_dict:
+ counter += 1
+ inter_layer_dict[i] = cell_number_max + counter
+
+
+ for j in range(1024):
+ for i in range(1024):
+ nissl_watershed_corrected[k][i][j] = inter_layer_dict[nissl_watershed[k][i][j]]
+
+ return nissl_watershed_corrected