added illumination correction option

macsima2mc/CLI.py

@@ -29,6 +29,12 @@ def get_args():
                     help='string specifying the name of the reference marker'
                     )
 
+    parser.add_argument('-ic',
+                    '--illumination_correction',
+                    action='store_true',
+                    help='Applies illumination correction to all tiles, the illumination profiles are created with basicpy'
+                    )
+
 
     parser.add_argument('-he',
                     '--hi_exposure_only',

macsima2mc/illumination_corr.py

@@ -0,0 +1,24 @@
+from basicpy import BaSiC
+import numpy as np
+
+def indices_per_channel(total_imgs,no_of_channels):
+    #total_imgs in the stack
+    img_indices=[ list( range(ch,total_imgs,no_of_channels) ) for ch in range( 0, no_of_channels )   ]
+    return img_indices
+
+def extract_channel_imgs (stack,indices):
+    return stack[indices,:,:]
+
+def apply_corr(uncorr_stack,no_of_channels):
+    corr_stack=np.zeros( uncorr_stack.shape,dtype=uncorr_stack.dtype )
+    total_imgs=uncorr_stack.shape[0]
+    indices=indices_per_channel(total_imgs, no_of_channels)
+    basic = BaSiC(get_darkfield=False, smoothness_flatfield=1.0,fitting_mode = "approximate", sort_intensity = True) 
+    for ind_list in indices:
+        uncorr_imgs=extract_channel_imgs(uncorr_stack, ind_list)
+        basic.fit(uncorr_imgs)
+        ffp=basic.flatfield
+        corr_stack[ind_list,:,:]=np.uint16( np.clip( uncorr_imgs.astype(float)/ffp  ,0, 65535) )
+    return corr_stack
+
+

macsima2mc/macsima2mc.py

@@ -12,10 +12,11 @@ def main(args):
     input=args.input
     output=args.output
     ref=args.reference_marker
+    basicpy_corr=args.illumination_correction 
     cycle_info=tools.cycle_info( input , macsima_pattern(version=2),ref_marker= ref)
     cycle_info=tools.append_metadata( cycle_info )
     #cycle_info.to_csv( args.output / 'cycle_{c}_info.csv'.format(c=f'{6:03d}'), index=False )
-    output_dirs=tools.create_stack( cycle_info, output ,ref_marker=ref,hi_exp=args.hi_exposure_only )
+    output_dirs=tools.create_stack( cycle_info, output ,ref_marker=ref,hi_exp=args.hi_exposure_only,ill_corr=basicpy_corr)
     [mc_tools.write_markers_file(path)  for path in output_dirs]
 
 

macsima2mc/tools.py

@@ -6,6 +6,7 @@
 import numpy as np
 from pathlib import Path
 import ome_writer
+import illumination_corr
 
 
 def merge_dicts(list_of_dicts):
@@ -183,7 +184,7 @@ def select_by_exposure(list_indices,exp_index=4,target='max'):
 
     return selected_indices
 
-def create_stack(cycle_info_df,output_dir,ref_marker='DAPI',hi_exp=False,extended_outputs=False):
+def create_stack(cycle_info_df,output_dir,ref_marker='DAPI',hi_exp=False,ill_corr=False,extended_outputs=False):
 
     if extended_outputs:
         out=outputs_dic()
@@ -225,7 +226,15 @@ def create_stack(cycle_info_df,output_dir,ref_marker='DAPI',hi_exp=False,extende
                 stack[counter,:,:]=tifff.imread(Path(target_path))
                 counter+=1
         stack_name =cast_stack_name(frame.cycle.iloc[0],index,conformed_markers)
-        stack_file_path= stack_output_dir/ stack_name
+
+        if ill_corr:
+            tag='corr_'
+            no_of_channels=len(conformed_markers)
+            stack=illumination_corr.apply_corr(stack,no_of_channels)
+        else:
+            tag=''
+
+        stack_file_path= stack_output_dir/ '{prefix}{base}'.format(prefix=tag,base=stack_name)
 
         if extended_outputs:
             out['index'].append(index)