Added fast autofocus integrated with Z stacking in the scan code
This commit is contained in:
Richard Bowman
parent
1cf1e40c54
commit
7e83d724b8
2 changed files with 71 additions and 19 deletions
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@ -19,6 +19,7 @@ class TileScanAPI(MicroscopeViewPlugin):
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style = payload.param('style', default='raster', convert=str)
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autofocus_dz = payload.param('autofocus_dz', default=50, convert=int)
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fast_autofocus = payload.param('fast_autofocus', default=False, convert=bool)
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use_video_port = payload.param('use_video_port', default=True, convert=bool)
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resize = payload.param('size', default=None)
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@ -43,6 +44,7 @@ class TileScanAPI(MicroscopeViewPlugin):
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use_video_port=use_video_port,
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resize=resize,
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bayer=bayer,
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fast_autofocus=fast_autofocus,
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metadata=metadata,
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tags=tags
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)
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@ -101,6 +101,7 @@ class ScanPlugin(MicroscopePlugin):
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use_video_port: bool = False,
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resize: Tuple[int, int] = None,
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bayer: bool = False,
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fast_autofocus = False,
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metadata: dict = {},
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tags: list = []):
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@ -124,6 +125,12 @@ class ScanPlugin(MicroscopePlugin):
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else:
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autofocus_enabled = False
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if fast_autofocus and not hasattr(self.microscope.plugin, 'default_fast_autofocus'):
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logging.error("Can't use fast autofocus in the scan - the plugin is missing or disabled.")
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fast_autofocus = False
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z_stack_dz = grid[2] * step_size[2] if grid[2] > 1 else 0 # shorthand for Z stack range
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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.
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# Construct an x-y grid (worry about z later)
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x_y_grid = construct_grid(
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initial_position,
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@ -132,27 +139,62 @@ class ScanPlugin(MicroscopePlugin):
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style=style
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)
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# Keep the initial Z position the same as our current position
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next_z = initial_position[2]
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if fast_autofocus: # If fast autofocus is enabled, make
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next_z += autofocus_dz/2 # sure we start from the top of the range
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initial_z = next_z # Save this value for use in raster scans
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# Now step through each point in the x-y coordinate array
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for line in x_y_grid:
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# If rastering, rather than snake (or eventually spiral)
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# Return focus to initial position
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if style == 'raster':
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# Return focus to initial position
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next_z = initial_z
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logging.debug("Returning to initial z position")
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self.microscope.stage.move_abs([line[0][0], line[0][1], initial_position[2]])
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self.microscope.stage.move_abs([line[0][0], line[0][1], next_z]) #RWB: I think this line is redundant
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for x_y in line:
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current_position = self.microscope.stage.position
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# Move to new grid position without changing z
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logging.debug("Moving to step {}".format([x_y[0], x_y[1], current_position[2]]))
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self.microscope.stage.move_abs([x_y[0], x_y[1], current_position[2]])
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logging.debug("Moving to step {}".format([x_y[0], x_y[1], next_z]))
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self.microscope.stage.move_abs([x_y[0], x_y[1], next_z])
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# Refocus
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if autofocus_enabled:
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# TODO: Better autofocus
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logging.debug("Running autofocus")
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self.microscope.plugin.default_autofocus.autofocus(
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range(-3 * autofocus_dz, 4 * autofocus_dz, autofocus_dz))
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logging.debug("Finished autofocus")
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time.sleep(1) # TODO: Remove
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if fast_autofocus:
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# TODO: put this in the fast autofocus plugin!
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with self.microscope.plugin.default_fast_autofocus.monitor_sharpness() as m:
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df = autofocus_dz
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# move down
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i, z = m.focus_rel(-df)
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# now inspect where the sharpest point is, and estimate the sharpness
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# (JPEG size) that we should find at the start of the Z stack
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jt, jz, js = m.move_data(i)
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best_z = jz[np.argmax(js)]
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target_s = np.interp([best_z+z_stack_dz/2.0], jz[::-1], js[::-1]) #NB jz is decreasing
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# now move to the start of the z stack
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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
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# We've deliberately undershot - figure out how much further we should move based on the curve
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current_js = m.jpeg_size()
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imax = np.argmax(js) # we want to crop out just the bit below the peak
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js = js[imax:] # NB z is in DECREASING order
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jz = jz[imax:]
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inow = np.argmax(js < current_js) # use the curve we recorded to estimate our position
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# TODO: fancy interpolation stuff
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# So, the Z position corresponding to our current sharpness value is zs[inow]
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# That means we should move forwards, by best_z - zs[inow]
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correction_move = best_z - jz[inow] - z_stack_dz/2.0
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logging.debug("Fast autofocus scan: correcting backlash by moving {} steps".format(correction_move))
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m.focus_rel(correction_move)
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else:
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logging.debug("Running autofocus")
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self.microscope.plugin.default_autofocus.autofocus(
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range(-3 * autofocus_dz, 4 * autofocus_dz, autofocus_dz))
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logging.debug("Finished autofocus")
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time.sleep(1) # TODO: Remove
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# If we're not doing a z-stack, just capture
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if (grid[2] <= 1):
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@ -172,13 +214,20 @@ class ScanPlugin(MicroscopePlugin):
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scan_id=scan_id,
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step_size=step_size[2],
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steps=grid[2],
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center=True,
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center=not fast_autofocus, # fast_autofocus does this for us!
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return_to_start=not fast_autofocus,
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use_video_port=use_video_port,
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resize=resize,
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bayer=bayer,
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metadata=metadata,
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tags=tags
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)
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# Make sure we use our current best estimate of focus (i.e. the current position) next point
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next_z = self.microscope.stage.position[2]
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if fast_autofocus:
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next_z += autofocus_dz/2 # Fast autofocus requires us to start at the top of the range
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if grid[2] > 1:
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next_z -= int(grid[2]/2.0*step_size[2]) # Z stacking means we're higher up to start with
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logging.debug("Returning to {}".format(initial_position))
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self.microscope.stage.move_abs(initial_position)
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@ -190,6 +239,7 @@ class ScanPlugin(MicroscopePlugin):
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step_size: int = 100,
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steps: int = 5,
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center: bool = True,
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return_to_start: bool = True,
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use_video_port: bool = False,
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resize: Tuple[int, int] = None,
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bayer: bool = False,
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@ -211,12 +261,12 @@ class ScanPlugin(MicroscopePlugin):
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# Store initial position
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initial_position = self.microscope.stage.position
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# Move to center scan
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if center:
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logging.debug("Moving to starting position")
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self.microscope.stage.move_rel([0, 0, int((-step_size * steps) / 2)])
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with self.microscope.lock:
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# Move to center scan
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if center:
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logging.debug("Moving to starting position")
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self.microscope.stage.move_rel([0, 0, int((-step_size * steps) / 2)])
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for i in range(steps):
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time.sleep(0.1)
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@ -234,6 +284,6 @@ class ScanPlugin(MicroscopePlugin):
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if i != steps - 1:
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logging.debug("Moving z by {}".format(step_size))
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self.microscope.stage.move_rel([0, 0, step_size])
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logging.debug("Returning to {}".format(initial_position))
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self.microscope.stage.move_abs(initial_position)
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if return_to_start:
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logging.debug("Returning to {}".format(initial_position))
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self.microscope.stage.move_abs(initial_position)
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