Added fast autofocus integrated with Z stacking in the scan code (7e83d724) · Commits · OpenFlexure / openflexure-microscope-server

openflexure_microscope/plugins/default/scan/api.py

+2 −0

Original line number	Diff line number	Diff line
		@@ -19,6 +19,7 @@ class TileScanAPI(MicroscopeViewPlugin):

		style = payload.param('style', default='raster', convert=str)
		autofocus_dz = payload.param('autofocus_dz', default=50, convert=int)
		fast_autofocus = payload.param('fast_autofocus', default=False, convert=bool)

		use_video_port = payload.param('use_video_port', default=True, convert=bool)
		resize = payload.param('size', default=None)
		@@ -43,6 +44,7 @@ class TileScanAPI(MicroscopeViewPlugin):
		use_video_port=use_video_port,
		resize=resize,
		bayer=bayer,
		fast_autofocus=fast_autofocus,
		metadata=metadata,
		tags=tags
		)

openflexure_microscope/plugins/default/scan/plugin.py

+69 −19

Original line number	Diff line number	Diff line
		@@ -101,6 +101,7 @@ class ScanPlugin(MicroscopePlugin):
		use_video_port: bool = False,
		resize: Tuple[int, int] = None,
		bayer: bool = False,
		fast_autofocus = False,
		metadata: dict = {},
		tags: list = []):

		@@ -124,6 +125,12 @@ class ScanPlugin(MicroscopePlugin):
		else:
		autofocus_enabled = False

		if fast_autofocus and not hasattr(self.microscope.plugin, 'default_fast_autofocus'):
		logging.error("Can't use fast autofocus in the scan - the plugin is missing or disabled.")
		fast_autofocus = False
		z_stack_dz = grid[2] * step_size[2] if grid[2] > 1 else 0 # shorthand for Z stack range
		sweep_to_scan_offset = 50 #TODO: make this a parameter, or calibrate it better! too small isn't a big problem, too big causes issues.

		# Construct an x-y grid (worry about z later)
		x_y_grid = construct_grid(
		initial_position,
		@@ -132,22 +139,57 @@ class ScanPlugin(MicroscopePlugin):
		style=style
		)

		# Keep the initial Z position the same as our current position
		next_z = initial_position[2]
		if fast_autofocus: # If fast autofocus is enabled, make
		next_z += autofocus_dz/2 # sure we start from the top of the range
		initial_z = next_z # Save this value for use in raster scans

		# Now step through each point in the x-y coordinate array
		for line in x_y_grid:
		# If rastering, rather than snake (or eventually spiral)
		if style == 'raster':
		# Return focus to initial position
		if style == 'raster':
		next_z = initial_z
		logging.debug("Returning to initial z position")
		self.microscope.stage.move_abs([line[0][0], line[0][1], initial_position[2]])
		self.microscope.stage.move_abs([line[0][0], line[0][1], next_z]) #RWB: I think this line is redundant

		for x_y in line:
		current_position = self.microscope.stage.position
		# Move to new grid position without changing z
		logging.debug("Moving to step {}".format([x_y[0], x_y[1], current_position[2]]))
		self.microscope.stage.move_abs([x_y[0], x_y[1], current_position[2]])
		logging.debug("Moving to step {}".format([x_y[0], x_y[1], next_z]))
		self.microscope.stage.move_abs([x_y[0], x_y[1], next_z])
		# Refocus
		if autofocus_enabled:
		# TODO: Better autofocus
		if fast_autofocus:
		# TODO: put this in the fast autofocus plugin!
		with self.microscope.plugin.default_fast_autofocus.monitor_sharpness() as m:
		df = autofocus_dz
		# move down
		i, z = m.focus_rel(-df)
		# now inspect where the sharpest point is, and estimate the sharpness
		# (JPEG size) that we should find at the start of the Z stack
		jt, jz, js = m.move_data(i)
		best_z = jz[np.argmax(js)]
		target_s = np.interp([best_z+z_stack_dz/2.0], jz[::-1], js[::-1]) #NB jz is decreasing

		# now move to the start of the z stack
		i, z = m.focus_rel(best_z - z + z_stack_dz/2.0 + sweep_to_scan_offset) # takes us to the start of the stack

		# We've deliberately undershot - figure out how much further we should move based on the curve
		current_js = m.jpeg_size()
		imax = np.argmax(js) # we want to crop out just the bit below the peak
		js = js[imax:] # NB z is in DECREASING order
		jz = jz[imax:]
		inow = np.argmax(js < current_js) # use the curve we recorded to estimate our position
		# TODO: fancy interpolation stuff

		# So, the Z position corresponding to our current sharpness value is zs[inow]
		# That means we should move forwards, by best_z - zs[inow]
		correction_move = best_z - jz[inow] - z_stack_dz/2.0
		logging.debug("Fast autofocus scan: correcting backlash by moving {} steps".format(correction_move))
		m.focus_rel(correction_move)

		else:
		logging.debug("Running autofocus")
		self.microscope.plugin.default_autofocus.autofocus(
		range(-3 * autofocus_dz, 4 * autofocus_dz, autofocus_dz))
		@@ -172,13 +214,20 @@ class ScanPlugin(MicroscopePlugin):
		scan_id=scan_id,
		step_size=step_size[2],
		steps=grid[2],
		center=True,
		center=not fast_autofocus, # fast_autofocus does this for us!
		return_to_start=not fast_autofocus,
		use_video_port=use_video_port,
		resize=resize,
		bayer=bayer,
		metadata=metadata,
		tags=tags
		)
		# Make sure we use our current best estimate of focus (i.e. the current position) next point
		next_z = self.microscope.stage.position[2]
		if fast_autofocus:
		next_z += autofocus_dz/2 # Fast autofocus requires us to start at the top of the range
		if grid[2] > 1:
		next_z -= int(grid[2]/2.0*step_size[2]) # Z stacking means we're higher up to start with

		logging.debug("Returning to {}".format(initial_position))
		self.microscope.stage.move_abs(initial_position)
		@@ -190,6 +239,7 @@ class ScanPlugin(MicroscopePlugin):
		step_size: int = 100,
		steps: int = 5,
		center: bool = True,
		return_to_start: bool = True,
		use_video_port: bool = False,
		resize: Tuple[int, int] = None,
		bayer: bool = False,
		@@ -211,13 +261,13 @@ class ScanPlugin(MicroscopePlugin):
		# Store initial position
		initial_position = self.microscope.stage.position


		with self.microscope.lock:
		# Move to center scan
		if center:
		logging.debug("Moving to starting position")
		self.microscope.stage.move_rel([0, 0, int((-step_size * steps) / 2)])

		with self.microscope.lock:

		for i in range(steps):
		time.sleep(0.1)
		logging.debug("Capturing...")
		@@ -234,6 +284,6 @@ class ScanPlugin(MicroscopePlugin):
		if i != steps - 1:
		logging.debug("Moving z by {}".format(step_size))
		self.microscope.stage.move_rel([0, 0, step_size])

		if return_to_start:
		logging.debug("Returning to {}".format(initial_position))
		self.microscope.stage.move_abs(initial_position)