def legacy():
"""
This is the code that I had originall wrote to do this.
This code was really messy, but worked, so I am leaving it here.
It has been rewritten in the above functions, but could be useful to look
at.
"""
## initialize the scope
scope=DGLab_Leica2.DGLab_Leica2()
## set up the
rows=string.ascii_uppercase[1:7]
columns=range(2,12)
sites=range(0,225)
pos_string=open("PositionList_96well_15x15sites.pos").read()
positions=StagePositions.positionList_parser(pos_string)
scope.mmc.enableContinuousFocus(True)
scope.set_illumination(Illuminations.Cy5)
#start_time=time.asctime().replace(" ","_").replace(":","_")
start_time="Thu_Aug_13_14_38_51_2015"
done_list={}
well_counter={}
## if this is a continuation of an old scan, this will add on to the
## existing log file.
if os.path.exists("Scan_%s.txt" % start_time):
completed_fin=open("Scan_%s.txt" % start_time,"r")
for line in completed_fin:
site=line.split(";")[0]
try:
if "No cells of interest" not in line.split(";")[1]:
well=site.split("-")
if well not in well_counter:
well_counter[well]=0
well_counter+=1
except:
pass
completed_fin.close()
status_out=open("Scan_%s.txt" % start_time,"r")
status_string=status_out.read()
status_out.close()
status_out=open("Scan_%s.txt" % start_time,"w")
status_out.write(status_string)
else:
status_out=open("Scan_%s.txt" % start_time,"w")
data_dir="Scan_%s" % start_time
if os.path.isdir(data_dir) is False:
os.mkdir(data_dir)
order=-1
for row in rows:
## flip the order that the columns are gone through
order=order*-1
for column in columns[::order]:
well="%s%s" % (row,column)
if well not in well_counter:well_counter[well]=0
if well_counter[well]<15:
## reset the well count for this new well.
well_count=well_counter[well]
## iterate through the site in this well.
for site in sites: ## Test case
if "%s%s-Site_%s" % (row,column,site) not in done_list.keys():
done_list["%s%s-Site_%s" % (row,column,site)]=True
print ">Looking at %s%s-Site_%s" % (row,column,site)
x,y=positions["%s%s-Site_%s" % (row,column,site)].get_xy()
## take an image to use for a trigger
scope.set_illumination(Illuminations.qCy5)
scope.mmc.setXYPosition("XYStage",x,y)
scope.mmc.snapImage()
img=scope.mmc.getImage()
## show image
plt.imshow(img)
plt.draw()
## analyze this image
cois=Threshold.coi_finder(img,
threshold=1000,
min_area=100,
max_area=1000)
if cois:
do_stack=True
progress_line=">%s%s-Site_%s;cois=%s\n" % (row,column,site,cois)
else:
do_stack=False
progress_line=">%s%s-Site_%s;No cells of interest\n" % (row,column,site)
status_out.write(progress_line)
## run the stack if there was a COI found
if do_stack:
print ">Stack at %s%s-Site_%s" % (row,column,site)
well_count+=1
well_counter[well]+=1
channels=[Illuminations.DIC,
Illuminations.GFP,
Illuminations.qCy5,
Illuminations.DAPI]
scope.mmc.enableContinuousFocus(False)
mda,md=scope.multi_channel_acquisition(channels)
FileOut.mda_save(out_dir=data_dir,
prefix="%s%s-Site_%s" % (row,column,site),
mda=mda,
md=md,
channels=channels)
print "Stack taken at %s%s-Site_%s" % (row,column,site)
## turn AFC back on
scope.mmc.enableContinuousFocus(True)
## move onto the next well if we have acquired enough
if well_count > 15:
break
## break out of the well if you can't focus
if scope.mmc.isContinuousFocusLocked() is False:
time.sleep(5)
if scope.mmc.isContinuousFocusLocked() is False:
print "Couldnt focus 3"
break
## break out of the column if you still cant focus
if scope.mmc.isContinuousFocusLocked() is False:
time.sleep(5)
if scope.mmc.isContinuousFocusLocked() is False:
print "Couldnt focus 2"
break
## break out of the row if you STILL cant focus.
if scope.mmc.isContinuousFocusLocked() is False:
time.sleep(5)
if scope.mmc.isContinuousFocusLocked() is False:
print "Couldnt focus 1"
break
status_out.close()
def plateScanSite(scope,
illumination,
mda_channels,
x,
y,
visible=True,
name="",
stack_height=5,
slice_height=.5,
minAreaCOI=100,
maxAreaCOI=1000,
thresholdCOI=1000):
"""
This is a function to run at each site of a 96well plate scan.
This requires:
-scope
-scan illumination
-MDA illuminations
-x
-y
Optional:
-visibility of scan image during acquisition
-name this is mostly for progress and debugging purposes
This returns:
-site_imaged: 0 or 1 depending if a MDA was taken
-MDA
-MDA_md
"""
scope.set_illumination(illumination)
scope.mmc.setXYPosition("XYStage",x,y)
scope.mmc.snapImage()
img=scope.mmc.getImage()
## show image
if visible is True:
plt.imshow(img)
plt.show()
plt.draw()
# analyze this image
cois=Threshold.coi_finder(img,
threshold=thresholdCOI,
min_area=minAreaCOI,
max_area=maxAreaCOI)
if cois:
do_stack=True
site_imaged=1
print "COIs found in %s, taking a stack" % name
else:
do_stack=False
site_imaged=0
md=None
mda=None
print "No COIs found in %s" % name
if do_stack:
scope.mmc.enableContinuousFocus(False)
mda,md=scope.multi_channel_acquisition(mda_channels,
z_width=stack_height,
z_increments=slice_height)
scope.mmc.enableContinuousFocus(True)
if visible is True: plt.close()
del img
return site_imaged, mda, md
