Update shutter behavior and LF acquisition channel order

.github/workflows/pytests.yml

@@ -11,7 +11,7 @@ jobs:
  runs-on: ubuntu-latest
  strategy:
  matrix:
- python-version: ['3.7', '3.8', '3.9', '3.10']
+ python-version: ['3.8', '3.9', '3.10']
 
  steps:
  - name: Checkout repo

.github/workflows/test_and_deploy.yml

@@ -21,7 +21,7 @@ jobs:
  strategy:
  matrix:
  platform: [ubuntu-latest, windows-latest, macos-latest]
- python-version: [3.7, 3.8, 3.9]
+ python-version: ['3.8', '3.9', '3.10']
 
  steps:
  - uses: actions/checkout@v2

README.md

@@ -30,9 +30,9 @@ The acquisition, calibration, background correction, reconstruction, and applica
 conda create -y -n recOrder python=3.9
 conda activate recOrder
 ```
-Install `napari` and `recOrder-napari`:
+Install `recOrder-napari`:
 ```
-pip install "napari[all]" recOrder-napari
+pip install recOrder-napari
 ```
 Open `napari` with `recOrder-napari`:
 ```

docs/microscope-installation-guide.md

@@ -15,9 +15,9 @@ conda create -y -n recOrder python=3.9
 conda activate recOrder
 ```
 
-Install `napari` and `recOrder`:
+Install `recOrder`:
 ```
-pip install "napari[all]" recOrder-napari
+pip install recOrder-napari
 ```
 Check your installation:
 ```

docs/software-installation-guide.md

@@ -7,9 +7,9 @@
 conda create -y -n recOrder python=3.9
 conda activate recOrder
 ```
-Install `napari` and `recOrder-napari`:
+Install `recOrder-napari`:
 ```
-pip install "napari[all]" recOrder-napari
+pip install recOrder-napari
 ```
 Open `napari` with `recOrder-napari`:
 ```

recOrder/acq/acq_functions.py

@@ -7,7 +7,7 @@
 import glob
 
 def generate_acq_settings(mm, channel_group, channels=None, zstart=None, zend=None, zstep=None,
- save_dir=None, prefix=None, keep_shutter_open = False):
+ save_dir=None, prefix=None, keep_shutter_open_channels=False, keep_shutter_open_slices=False):
  """
  This function generates a json file specific to the micromanager SequenceSettings.
  It has default parameters for a multi-channels z-stack acquisition but does not yet
@@ -74,7 +74,8 @@ def generate_acq_settings(mm, channel_group, channels=None, zstart=None, zend=No
  original_json['relativeZSlice'] = True
  original_json['slicesFirst'] = True
  original_json['timeFirst'] = False
- original_json['keepShutterOpenSlices'] = keep_shutter_open
+ original_json['keepShutterOpenSlices'] = keep_shutter_open_slices
+ original_json['keepShutterOpenChannels'] = keep_shutter_open_channels
  original_json['useAutofocus'] = False
  original_json['saveMode'] = 'MULTIPAGE_TIFF'
  original_json['save'] = True if save_dir else False

recOrder/calib/Calibration.py

@@ -7,6 +7,7 @@
  set_lc_state, snap_and_average, snap_and_get_image, get_lc, define_config_state
 from recOrder.calib.Optimization import BrentOptimizer, MinScalarOptimizer
 from mpl_toolkits.axes_grid1.axes_divider import make_axes_locatable
+from napari.utils.notifications import show_warning
 from scipy.interpolate import interp1d
 from scipy.stats import linregress
 from scipy.optimize import least_squares
@@ -145,6 +146,11 @@ def __init__(self, mmc, mm, group='Channel', lc_control_mode='MM-Retardance', in
  self.directory = None
  self.inst_mat = None
 
+ # Shutter
+ self.shutter_device = self.mmc.getShutterDevice()
+ self._auto_shutter_state = None
+ self._shutter_state = None
+
  def set_dacs(self, lca_dac, lcb_dac):
  self.PROPERTIES['LCA-DAC'] = (f'TS_{lca_dac}', 'Volts')
  self.PROPERTIES['LCB-DAC'] = (f'TS_{lcb_dac}', 'Volts')
@@ -639,13 +645,46 @@ def opt_I135(self, lca_bound, lcb_bound):
  logging.info("--------done--------")
  logging.debug("--------done--------")
 
- def calc_blacklevel(self):
+ def open_shutter(self):
+ if self.shutter_device == '': # no shutter
+ input('Please manually open the shutter and press <Enter>')
+ else:
+ self.mmc.setShutterOpen(True)
+
+ def reset_shutter(self):
+ """
+ Return autoshutter to its original state before closing
 
- auto_shutter = self.mmc.getAutoShutter()
- shutter = self.mmc.getShutterOpen()
+ Returns
+ -------
 
- self.mmc.setAutoShutter(False)
- self.mmc.setShutterOpen(False)
+ """
+ if self.shutter_device == '': # no shutter
+ input('Please reset the shutter to its original state and press <Enter>')
+ logging.info("This is the end of the command-line instructions. You can return to the napari window.")
+ else:
+ self.mmc.setAutoShutter(self._auto_shutter_state)
+ self.mmc.setShutterOpen(self._shutter_state)
+
+ def close_shutter_and_calc_blacklevel(self):
+ self._auto_shutter_state = self.mmc.getAutoShutter()
+ self._shutter_state = self.mmc.getShutterOpen()
+
+ if self.shutter_device == '': # no shutter
+ show_warning('No shutter found. Please follow the command-line instructions...')
+ shutter_warning_msg = """
+ recOrder could not find an automatic shutter configured through Micro-Manager.
+ >>> If you would like manually enter the black level, enter an integer or float and press <Enter>
+ >>> If you would like to estimate the black level, please close the shutter and press <Enter> 
+ """
+
+ in_string = input(shutter_warning_msg)
+ if in_string.isdigit(): # True if positive integer
+ self.I_Black = float(in_string)
+ return
+ else:
+ self.mmc.setAutoShutter(False)
+ self.mmc.setShutterOpen(False)
 
  n_avg = 20
  avgs = []
@@ -655,16 +694,8 @@ def calc_blacklevel(self):
  avgs.append(mean)
 
  blacklevel = np.mean(avgs)
-
- self.mmc.setAutoShutter(auto_shutter)
-
- if not auto_shutter:
- self.mmc.setShutterOpen(shutter)
-
  self.I_Black = blacklevel
 
- return blacklevel
-
  def get_full_roi(self):
  # Get Image Parameters
  self.mmc.snapImage()
@@ -718,131 +749,6 @@ def display_and_check_ROI(self, rect):
  else:
  raise ValueError('Did not understand your answer, please check spelling')
 
- def run_5state_calibration(self, param):
- """
- Param is a list or tuple of:
- (swing, wavelength, lc_bounds, black level)
- """
- self.swing = param[0]
- self.wavelength = param[1]
- self.meta_file = param[2]
- use_full_FOV = param[3]
-
- # Get Image Parameters
- self.mmc.snapImage()
- self.mmc.getImage()
- self.height, self.width = self.mmc.getImageHeight(), self.mmc.getImageWidth()
- self.ROI = (0, 0, self.width, self.height)
-
- # Check if change of ROI is needed
- if use_full_FOV is False:
- rect = self.check_and_get_roi()
- cont = self.display_and_check_ROI(rect)
-
- if not cont:
- print('\n---------Stopping Calibration---------\n')
- return
- else:
- self.mmc.setROI(rect.x, rect.y, rect.width, rect.height)
- self.ROI = (rect.x, rect.y, rect.width, rect.height)
-
- # Calculate Blacklevel
- logging.debug('Calculating Blacklevel ...')
- self.I_Black = self.calc_blacklevel()
- logging.debug(f'Blacklevel: {self.I_Black}\n')
-
- # Set LC Wavelength:
- if self.mode == 'MM-Retardance':
- self.mmc.setProperty(LC_DEVICE_NAME, 'Wavelength', self.wavelength)
-
- self.opt_Iext()
- self.opt_I0()
- self.opt_I45(0.05, 0.05)
- self.opt_I90(0.05, 0.05)
- self.opt_I135(0.05, 0.05)
-
- # Calculate Extinction
- self.extinction_ratio = self.calculate_extinction()
-
- # Write Metadata
- self.write_metadata()
-
- # Return ROI to full FOV
- if use_full_FOV is False:
- self.mmc.clearROI()
-
- logging.info("\n=======Finished Calibration=======\n")
- logging.info(f"EXTINCTION = {self.extinction_ratio}")
- logging.debug("\n=======Finished Calibration=======\n")
- logging.debug(f"EXTINCTION = {self.extinction_ratio}")
-
- def run_4state_calibration(self, param):
- """
- Param is a list or tuple of:
- (swing, wavelength, lc_bounds, black level, <mode>)
- where <mode> is one of 'full','coarse','fine'
- """
- self.swing = param[0]
- self.wavelength = param[1]
- self.meta_file = param[2]
- use_full_FOV = param[3]
-
- # Get Image Parameters
- self.mmc.snapImage()
- self.mmc.getImage()
- self.height, self.width = self.mmc.getImageHeight(), self.mmc.getImageWidth()
- self.ROI = (0, 0, self.width, self.height)
-
- # Check if change of ROI is needed
- if use_full_FOV is False:
- rect = self.check_and_get_roi()
- cont = self.display_and_check_ROI(rect)
-
- if not cont:
- print('\n---------Stopping Calibration---------\n')
- return
- else:
- self.mmc.setROI(rect.x, rect.y, rect.width, rect.height)
- self.ROI = (rect.x, rect.y, rect.width, rect.height)
-
- # Calculate Blacklevel
- print('Calculating Blacklevel ...')
- self.I_Black = self.calc_blacklevel()
- print(f'Blacklevel: {self.I_Black}\n')
-
- # Set LC Wavelength:
- if self.mode == 'MM-Retardance':
- self.mmc.setProperty(LC_DEVICE_NAME, 'Wavelength', self.wavelength)
-
- self.opt_Iext()
- self.opt_I0()
- self.opt_I60(0.05, 0.05)
- self.opt_I120(0.05, 0.05)
-
- # Calculate Extinction
- self.extinction_ratio = self.calculate_extinction()
-
- # Write Metadata
- self.write_metadata()
-
- # Return ROI to full FOV
- if use_full_FOV is False:
- self.mmc.clearROI()
-
- print("\n=======Finished Calibration=======\n")
- print(f"EXTINCTION = {self.extinction_ratio}")
-
- def run_calibration(self, scheme, options):
-
- if scheme == '5-State':
- self.run_5state_calibration(options)
-
- elif scheme == '4-State Extinction':
- self.run_4state_calibration(options)
-
- else:
- raise ValueError('Please define the calibration scheme')
-
  def calculate_extinction(self, swing, black_level, intensity_extinction, intensity_elliptical):
  return np.round((1 / np.sin(np.pi * swing) ** 2) * \
  (intensity_elliptical - black_level) / (intensity_extinction - black_level), 2)

recOrder/plugin/widget/main_widget.py

@@ -2417,7 +2417,8 @@ def load_calibration(self):
  self.swing = metadata.Swing
 
  # Initialize calibration class
- self.calib = QLIPP_Calibration(self.mmc, self.mm, group=self.config_group)
+ self.calib = QLIPP_Calibration(self.mmc, self.mm, group=self.config_group, lc_control_mode=self.calib_mode,
+ interp_method=self.interp_method, wavelength=self.wavelength)
  self.calib.swing = self.swing
  self.ui.le_swing.setText(str(self.swing))
  self.calib.wavelength = self.wavelength

recOrder/plugin/workers/acquisition_workers.py

@@ -855,7 +855,8 @@ def work(self):
  channel_group=self.channel_group,
  channels=channels,
  save_dir=self.snap_dir,
- prefix=self.prefix)
+ prefix=self.prefix,
+ keep_shutter_open_channels=True)
  self._check_abort()
  # acquire images
  stack = self._acquire()
@@ -872,9 +873,16 @@ def work(self):
  zend=self.calib_window.z_end,
  zstep=self.calib_window.z_step,
  save_dir=self.snap_dir,
- prefix=self.prefix)
+ prefix=self.prefix,
+ keep_shutter_open_channels=True,
+ keep_shutter_open_slices=True)
 
  self._check_abort()
+
+ # set acquisition order to channel-first
+ self.settings['slicesFirst'] = False
+ self.settings['acqOrderMode'] = 0 # TIME_POS_SLICE_CHANNEL
+
  # acquire images
  stack = self._acquire()
 

recOrder/plugin/workers/calibration_workers.py

@@ -70,16 +70,20 @@ def work(self):
 
  self._check_abort()
 
- # Calculate Blacklevel
  logging.info('Calculating Black Level ...')
  logging.debug('Calculating Black Level ...')
- self.calib.calc_blacklevel()
+ self.calib.close_shutter_and_calc_blacklevel()
+
+ # Calculate Blacklevel
  logging.info(f'Black Level: {self.calib.I_Black:.0f}\n')
  logging.debug(f'Black Level: {self.calib.I_Black:.0f}\n')
 
  self._check_abort()
  self.progress_update.emit((10, 'Calibrating Extinction State...'))
 
+ # Open shutter
+ self.calib.open_shutter()
+
  # Set LC Wavelength:
  self.calib.set_wavelength(int(self.calib_window.wavelength))
  if self.calib_window.calib_mode == 'MM-Retardance':
@@ -90,6 +94,9 @@ def work(self):
  # Optimize States
  self._calibrate_4state() if self.calib_window.calib_scheme == '4-State' else self._calibrate_5state()
 
+ # Reset shutter autoshutter
+ self.calib.reset_shutter()
+
  # Return ROI to full FOV
  if self.calib_window.use_cropped_roi:
  self.calib_window.mmc.clearROI()
@@ -356,11 +363,13 @@ def load_calibration(calib, metadata: MetadataReader):
 
  # Calculate black level after loading these properties
  calib.intensity_emitter = MockEmitter()
- calib.calc_blacklevel()
+ calib.close_shutter_and_calc_blacklevel()
+ calib.open_shutter()
  set_lc_state(calib.mmc, calib.group, 'State0')
  calib.I_Ext = snap_and_average(calib.snap_manager)
  set_lc_state(calib.mmc, calib.group, 'State1')
  calib.I_Elliptical = snap_and_average(calib.snap_manager)
+ calib.reset_shutter()
 
  yield str(calib.calculate_extinction(calib.swing, calib.I_Black, calib.I_Ext, calib.I_Elliptical))