small-fish-gui 1.3.5__py3-none-any.whl → 1.5.0__py3-none-any.whl

small_fish_gui/batch/values.txt

@@ -0,0 +1,65 @@
+List of keys for batch 'values' dict instance :
+
+Batch_folder 
+0
+image path 
+3D stack 
+multichannel 
+Dense regions deconvolution 
+Cluster computation 
+Segmentation 
+Napari correction 
+x 
+y 
+z 
+c 
+t 
+cyto_model_name 
+cytoplasm channel 
+cytoplasm diameter 
+nucleus_model_name 
+nucleus channel 
+nucleus diameter 
+Segment only nuclei 
+show segmentation 
+saving path 
+filename 
+threshold 
+threshold penalty 
+channel to compute 
+voxel_size_z 
+voxel_size_y 
+voxel_size_x 
+spot_size_z 
+spot_size_y 
+spot_size_x 
+log_kernel_size_z 
+log_kernel_size_y 
+log_kernel_size_x 
+minimum_distance_z 
+minimum_distance_y 
+minimum_distance_x 
+nucleus channel signal 
+alpha 
+beta 
+gamma 
+deconvolution_kernel_z 
+deconvolution_kernel_y 
+deconvolution_kernel_x 
+cluster size 
+min number of spots 
+Interactive threshold selector 
+spots_extraction_folder 
+spots_filename 
+do_spots_csv 
+do_spots_excel 
+do_spots_feather 
+output_folder 
+batch_name 
+save segmentation 
+save detection 
+extract spots 
+csv 
+xlsx 
+feather 
+2

small_fish_gui/docs/conf.py

@@ -0,0 +1 @@
+#init conf file

small_fish_gui/gui/animation.py

@@ -10,21 +10,30 @@ WAITING_TEXT = [
 
 def add_default_loading(funct) :
     def inner(*args,**kwargs) :
-        random_text = np.random.randint(0,len(WAITING_TEXT))
-        waiting_layout = [
-        [sg.Text(WAITING_TEXT[random_text], font= '10')]
-        ]
-        window = sg.Window(
-        title= 'small_fish',
-        layout= waiting_layout,
-        grab_anywhere= True,
-        finalize=True
-    )
-
-        window.read(timeout= 30, close= False)
-        try :
+
+        hide_loading = kwargs.get("hide_loading")
+        if 'hide_loading' in kwargs : del kwargs['hide_loading']
+
+        if not hide_loading :
+            random_text = np.random.randint(0,len(WAITING_TEXT))
+            waiting_layout = [
+            [sg.Text(WAITING_TEXT[random_text], font= '10')]
+            ]
+            window = sg.Window(
+            title= 'small_fish',
+            layout= waiting_layout,
+            grab_anywhere= True,
+            finalize=True
+        )
+
+            window.read(timeout= 30, close= False)
+            try :
+                return funct(*args, **kwargs)
+            finally :
+                window.close()
+
+        else : 
             return funct(*args, **kwargs)
-        finally :
-            window.close()
+
     return inner
 

small_fish_gui/gui/layout.py

@@ -95,7 +95,7 @@ def path_layout(keys= [],look_for_dir = False, header=None, preset=os.getcwd())
 
     max_length = len(max(keys, key=len))
     layout = [
-        [sg.Text(pad_right(name, max_length, ' ')), Browse(key= name, initial_folder= preset)] for name in keys
+        [sg.Text(pad_right(name, max_length, ' ')), Browse(key= name, target=(555666777,2), initial_folder= preset), sg.Text('')] for name in keys
         ]
     if isinstance(header, str) :
         layout = [add_header(header)] + layout
@@ -152,7 +152,20 @@ def radio_layout(values, header=None) :
         layout = [add_header(header)] + layout
     return layout
 
-def _segmentation_layout(multichannel, cytoplasm_model_preset= 'cyto2', nucleus_model_preset= 'nuclei', cytoplasm_channel_preset=0, nucleus_channel_preset=0, cyto_diameter_preset=30, nucleus_diameter_preset= 30, show_segmentation_preset= False, segment_only_nuclei_preset=False, saving_path_preset=os.getcwd(), filename_preset='cell_segmentation.png',) :
+def _segmentation_layout(
+        multichannel, 
+        cytoplasm_model_preset= 'cyto3', 
+        nucleus_model_preset= 'nuclei', 
+        cytoplasm_channel_preset=0, 
+        nucleus_channel_preset=0, 
+        other_nucleus_image_preset = None,
+        cyto_diameter_preset=30, 
+        nucleus_diameter_preset= 30, 
+        show_segmentation_preset= False, 
+        segment_only_nuclei_preset=False, 
+        saving_path_preset=os.getcwd(), 
+        filename_preset='cell_segmentation.png',
+        ) :
     
     USE_GPU = use_gpu()
 
@@ -160,7 +173,9 @@ def _segmentation_layout(multichannel, cytoplasm_model_preset= 'cyto2', nucleus_
     if len(models_list) == 0 : models_list = ['no model found']
     
     #Header : GPU availabality
-    layout = [[sg.Text("GPU is currently "), sg.Text('ON', text_color= 'green') if USE_GPU else sg.Text('OFF', text_color= 'red')]]
+    layout = [
+        [sg.Text("GPU is currently "), sg.Text('ON', text_color= 'green') if USE_GPU else sg.Text('OFF', text_color= 'red')]
+        ]
     
     #cytoplasm parameters
     layout += [
@@ -170,6 +185,8 @@ def _segmentation_layout(multichannel, cytoplasm_model_preset= 'cyto2', nucleus_
                         ]
                         
     if multichannel : layout += parameters_layout(['cytoplasm channel'],default_values= [cytoplasm_channel_preset])
+
+
     layout += parameters_layout(['cytoplasm diameter'], unit= "px", default_values= [cyto_diameter_preset])
     #Nucleus parameters
     layout += [
@@ -179,6 +196,7 @@ def _segmentation_layout(multichannel, cytoplasm_model_preset= 'cyto2', nucleus_
                 ]
     
     if multichannel : layout += parameters_layout(['nucleus channel'], default_values= [nucleus_channel_preset])
+    layout += path_layout(['other_nucleus_image'], preset=other_nucleus_image_preset)
     layout += parameters_layout([ 'nucleus diameter'],unit= "px", default_values= [nucleus_diameter_preset])
     layout += bool_layout(["Segment only nuclei"], preset=segment_only_nuclei_preset)
     
@@ -246,7 +264,12 @@ def _detection_layout(
     unit = {'voxel_size' : 'nm', 'minimum_distance' : 'nm', 'spot_size' : 'radius(nm)', 'log_kernel_size' : 'px'}
 
     layout += tuple_layout(opt=opt, unit=unit, default_dict=default_dict, voxel_size= tuple_shape, spot_size= tuple_shape, log_kernel_size= tuple_shape, minimum_distance= tuple_shape)
-
+    
+    if (do_segmentation and is_multichannel) or (is_multichannel and segmentation_done):
+        default_segmentation = [default_dict.setdefault('nucleus channel signal', default_dict.setdefault('nucleus channel',0))]
+        layout += [[sg.Text("nucleus channel signal "), sg.InputText(default_text=default_segmentation, key= "nucleus channel signal", size= 5, tooltip= "Channel from which signal will be measured for nucleus features, \nallowing you to measure signal from a different channel than the one used for segmentation.")]]
+        # layout += parameters_layout(['nucleus channel signal'], default_values=default_segmentation) + [[sg.Text("Channel from which signal will be measured for nucleus features, allowing you to measure signal from a different channel than the one used for segmentation.")]]
+    
     #Deconvolution
     if do_dense_region_deconvolution :
         default_dense_regions_deconvolution = [default_dict.setdefault('alpha',0.5), default_dict.setdefault('beta',1)]
@@ -259,9 +282,7 @@ def _detection_layout(
         layout += parameters_layout(['cluster size'], unit="radius(nm)", default_values=[default_dict.setdefault('cluster size',400)])
         layout += parameters_layout(['min number of spots'], default_values=[default_dict.setdefault('min number of spots', 5)])
 
-    if (do_segmentation and is_multichannel) or (is_multichannel and segmentation_done):
-        default_segmentation = [default_dict.setdefault('nucleus channel signal', default_dict.setdefault('nucleus channel',0))]
-        layout += parameters_layout(['nucleus channel signal'], default_values=default_segmentation) + [[sg.Text(" channel from which signal will be measured for nucleus features.")]]
+    
 
     layout += bool_layout(['Interactive threshold selector'], preset=[False])
     layout += path_layout(

small_fish_gui/gui/prompts.py

@@ -32,14 +32,14 @@ def prompt(layout, add_ok_cancel=True, timeout=None, timeout_key='TIMEOUT_KEY',
         window.close()
         return event, values
 
-def prompt_with_help(layout, help =None, add_scrollbar=True) :
+def prompt_with_help(layout, help =None, add_scrollbar=True, vertical_scroll_only=True) :
     layout += [[]]
     layout += [[sg.Button('Help')]]
     layout += [[sg.Button('Ok'), sg.Button('Cancel')]]
     
     if add_scrollbar :
         size = (400,500)
-        col_elmt = sg.Column(layout, scrollable=True, vertical_scroll_only=True, size=size)
+        col_elmt = sg.Column(layout, scrollable=True, vertical_scroll_only=vertical_scroll_only, size=size)
         layout = [[col_elmt]]
     else :
         size = (None,None)
@@ -280,11 +280,13 @@ def _warning_popup(warning:str) :
 
 def _sumup_df(results: pd.DataFrame) :
 
+    COLUMNS = ['acquisition_id','threshold', 'spot_number', 'cell_number', 'filename', 'channel to compute']
+
     if len(results) > 0 :
         if 'channel to compute' not in results : results['channel to compute'] = np.NaN
-        res = results.loc[:,['acquisition_id', 'spot_number', 'cell_number', 'filename', 'channel to compute']]
+        res = results.loc[:,COLUMNS]
     else :
-        res = pd.DataFrame(columns= ['acquisition_id', 'spot_number', 'cell_number', 'filename', 'channel to compute'])
+        res = pd.DataFrame(columns= COLUMNS)
 
     return res
 
@@ -293,14 +295,14 @@ def hub_prompt(fov_results, do_segmentation=False) :
     sumup_df = _sumup_df(fov_results)
     
     if do_segmentation :
-        segmentation_object = sg.Text('Segmentation was performed', font='8', text_color= 'green')
+        segmentation_object = sg.Text('Segmentation in memory', font='8', text_color= 'green')
     else :
-        segmentation_object = sg.Text('Segmentation was not performed', font='8', text_color= 'red')
+        segmentation_object = sg.Text('No segmentation in memory', font='8', text_color= 'red')
 
     layout = [
         [sg.Text('RESULTS', font= 'bold 13')],
         [sg.Table(values= list(sumup_df.values), headings= list(sumup_df.columns), row_height=20, num_rows= 5, vertical_scroll_only=False, key= "result_table"), segmentation_object],
-        [sg.Button('Add detection'), sg.Button('Compute colocalisation')],#, sg.Button('Batch detection')],
+        [sg.Button('Add detection'), sg.Button('Compute colocalisation'), sg.Button('Batch detection')],
         [sg.Button('Save results', button_color= 'green'), sg.Button('Delete acquisitions',button_color= 'gray'), sg.Button('Reset segmentation',button_color= 'gray'), sg.Button('Reset results',button_color= 'gray')]
         # [sg.Button('Save results', button_color= 'green'), sg.Button('Reset results',button_color= 'gray')]
     ]
@@ -333,7 +335,7 @@ def ask_detection_confirmation(used_threshold) :
         [sg.Button("Ok"), sg.Button("Restart detection")]
     ]
 
-    event, value = prompt(layout, add_ok_cancel=False)
+    event, value = prompt(layout, add_ok_cancel=False, add_scrollbar=False)
 
     if event == 'Restart detection' :
         return False
@@ -346,7 +348,7 @@ def ask_cancel_detection() :
         [sg.Button("Yes"), sg.Button("No")]
     ]
 
-    event, value = prompt(layout, add_ok_cancel=False)
+    event, value = prompt(layout, add_ok_cancel=False, add_scrollbar=False)
 
     if event == 'No' :
         return False

small_fish_gui/interface/output.py

@@ -10,9 +10,11 @@ def _cast_spot_to_tuple(spot) :
 def _cast_spots_to_tuple(spots) :
     return tuple(list(map(_cast_spot_to_tuple, spots)))
 
-def write_results(dataframe: pd.DataFrame, path:str, filename:str, do_excel= True, do_feather= False, do_csv=False) :
+def write_results(dataframe: pd.DataFrame, path:str, filename:str, do_excel= True, do_feather= False, do_csv=False, overwrite=False) :
     check_parameter(dataframe= pd.DataFrame, path= str, filename = str, do_excel = bool, do_feather = bool)
 
+    dataframe.columns = dataframe.columns.astype(str) # assert columns header are string for feather
+
     if len(dataframe) == 0 : return True
     if not do_excel and not do_feather and not do_csv : 
         return False
@@ -23,9 +25,11 @@ def write_results(dataframe: pd.DataFrame, path:str, filename:str, do_excel= Tru
 
     new_filename = filename
     i= 1
-    while new_filename + '.xlsx' in os.listdir(path) or new_filename + '.feather' in os.listdir(path) or new_filename + '.csv' in os.listdir(path) :
-        new_filename = filename + '_{0}'.format(i)
-        i+=1
+
+    if not overwrite :
+        while new_filename + '.xlsx' in os.listdir(path) or new_filename + '.feather' in os.listdir(path) or new_filename + '.csv' in os.listdir(path) :
+            new_filename = filename + '_{0}'.format(i)
+            i+=1
 
     if 'image' in dataframe.columns :
         dataframe = dataframe.drop(['image'], axis=1)

small_fish_gui/pipeline/__init__.py

@@ -0,0 +1,21 @@
+"""
+Module containing main pipeline for user mode as well as calls for pipeline functions.
+"""
+
+from ._preprocess import reorder_shape
+from ._preprocess import reorder_image_stack
+from ._preprocess import prepare_image_detection
+from ._preprocess import convert_parameters_types
+
+from ._segmentation import launch_segmentation
+from ._segmentation import _cast_segmentation_parameters
+from ._segmentation import cell_segmentation
+from ._segmentation import plot_segmentation
+
+from .detection import launch_detection
+from .detection import launch_features_computation
+from .detection import launch_cell_extraction
+from .detection import get_nucleus_signal
+from .detection import output_spot_tiffvisual
+
+from .spots import launch_spots_extraction

small_fish_gui/pipeline/_napari_wrapper.py

@@ -2,59 +2,26 @@
 Contains Napari wrappers to visualise and correct spots/clusters.
 """
 
+import napari.layers
+import napari.types
 import numpy as np
-import scipy.ndimage as ndi
 import napari
 
-from napari.utils.events import Event
-from napari.layers import Points
 from bigfish.stack import check_parameter
 from ..utils import compute_anisotropy_coef
 from ._colocalisation import spots_multicolocalisation
 
-class Points_callback :
-    """
-    Custom class to handle points number evolution during Napari run.
-    """
-    
-    def __init__(self, points, next_id) -> None:
-        self.points = points
-        self.next_id = next_id
-        self._set_callback()
-    
-    def __str__(self) -> str:
-        string = 'Points_callback object state :\ncurrent_points_number : {0}\ncurrnet_id : {1}'.format(self.current_points_number, self.next_id)
-        return string
-    
-    def get_points(self) :
-        return self.points
-    
-    def get_next_id(self) : 
-        return self.next_id
-    
-    def _set_callback(self) :
-        def callback(event:Event) :
-
-            old_points = self.get_points()
-            new_points:Points = event.source.data
-            features = event.source.features
-            
-            current_point_number = len(old_points)
-            next_id = self.get_next_id()
-            new_points_number = len(new_points)
-
-            if new_points_number > current_point_number :
-                features.at[new_points_number - 1, "id"] = next_id
-                self.next_id += 1
-
-            #preparing next callback
-            self.points = new_points
-            self._set_callback()
-        self.callback = callback
-
 def _update_clusters(new_clusters: np.ndarray, spots: np.ndarray, voxel_size, cluster_size, min_spot_number, shape) :
     if len(new_clusters) == 0 : return new_clusters
     if len(spots) == 0 : return new_clusters
+
+    if len(new_clusters[0]) in [2,3] :
+        new_clusters = np.concatenate([
+            new_clusters,
+            np.zeros(shape=(len(new_clusters),1), dtype=int),
+            np.arange(len(new_clusters), dtype=int).reshape(len(new_clusters),1)
+            ],axis=1, dtype=int)
+
     assert len(new_clusters[0]) == 4 or len(new_clusters[0]) == 5, "Wrong number of coordinates for clusters should not happen."
     
     # Update spots clusters
@@ -93,45 +60,48 @@ def correct_spots(image, spots, voxel_size= (1,1,1), clusters= None, cluster_siz
         )
 
     scale = compute_anisotropy_coef(voxel_size)
-    try :
-        Viewer = napari.Viewer(ndisplay=2, title= 'Spot correction', axis_labels=['z','y','x'], show= False)
-        Viewer.add_image(image, scale=scale, name= "rna signal", blending= 'additive', colormap='red', contrast_limits=[image.min(), image.max()])
-        other_colors = ['green', 'blue', 'gray', 'cyan', 'bop orange', 'bop purple'] * ((len(other_images)-1 // 7) + 1)
-        for im, color in zip(other_images, other_colors) : 
-            Viewer.add_image(im, scale=scale, blending='additive', visible=False, colormap=color, contrast_limits=[im.min(), im.max()])
-        layer_offset = len(other_images)
-
-        Viewer.add_points(spots, size = 5, scale=scale, face_color= 'green', opacity= 1, symbol= 'ring', name= 'single spots') # spots
-        if type(clusters) != type(None) : Viewer.add_points(clusters[:,:dim], size = 10, scale=scale, face_color= 'blue', opacity= 0.7, symbol= 'diamond', name= 'foci', features= {"spot_number" : clusters[:,dim], "id" : clusters[:,dim+1]}, feature_defaults= {"spot_number" : 0, "id" : -1}) # cluster
-        if type(cell_label) != type(None) and not np.array_equal(nucleus_label, cell_label) : Viewer.add_labels(cell_label, scale=scale, opacity= 0.2, blending= 'additive')
-        if type(nucleus_label) != type(None) : Viewer.add_labels(nucleus_label, scale=scale, opacity= 0.2, blending= 'additive')
-        
-        #prepare cluster update
-        if type(clusters) != type(None) : 
-            next_cluster_id = clusters[-1,-1] + 1 if len(clusters) > 0 else 1
-            _callback = Points_callback(points=clusters[:dim], next_id=next_cluster_id)
-            points_callback = Viewer.layers[2 + layer_offset].events.data.connect((_callback, 'callback'))
-        Viewer.show(block=False)
-        napari.run()
-        
-
-        new_spots = np.array(Viewer.layers[1 + layer_offset].data, dtype= int)
+    Viewer = napari.Viewer(ndisplay=2, title= 'Spot correction', axis_labels=['z','y','x'], show= False)
+    Viewer.add_image(image, scale=scale, name= "rna signal", blending= 'additive', colormap='red', contrast_limits=[image.min(), image.max()])
+    other_colors = ['green', 'blue', 'gray', 'cyan', 'bop orange', 'bop purple'] * ((len(other_images)-1 // 7) + 1)
+    for im, color in zip(other_images, other_colors) : 
+        Viewer.add_image(im, scale=scale, blending='additive', visible=False, colormap=color, contrast_limits=[im.min(), im.max()])
+    layer_offset = len(other_images)
+
+    Viewer.add_points(  # single molecule spots; this layer can be update by user.
+        spots, 
+        size = 5, 
+        scale=scale, 
+        face_color= 'transparent', 
+        opacity= 1, 
+        symbol= 'disc', 
+        name= 'single spots'
+        )
+    
+    if type(clusters) != type(None) : Viewer.add_points( # cluster; this layer can be update by user.
+        clusters[:,:dim], 
+        size = 10, 
+        scale=scale, 
+        face_color= 'blue', 
+        opacity= 0.7, 
+        symbol= 'diamond', 
+        name= 'foci', 
+        features= {"spot_number" : clusters[:,dim], "id" : clusters[:,dim+1]}, 
+        feature_defaults= {"spot_number" : 0, "id" : -1}
+        )
 
-        if type(clusters) != type(None) :
-            if len(clusters) > 0 : 
-                new_clusters = np.concatenate([
-                    np.array(Viewer.layers[2 + layer_offset].data, dtype= int),
-                    np.array(Viewer.layers[2 + layer_offset].features, dtype= int)
-                ],
-                axis= 1)
+    if type(cell_label) != type(None) and not np.array_equal(nucleus_label, cell_label) : Viewer.add_labels(cell_label, scale=scale, opacity= 0.2, blending= 'additive')
+    if type(nucleus_label) != type(None) : Viewer.add_labels(nucleus_label, scale=scale, opacity= 0.2, blending= 'additive')
+        
+    Viewer.show(block=False)
+    napari.run()
 
-                new_clusters = _update_clusters(new_clusters, new_spots, voxel_size=voxel_size, cluster_size=cluster_size, min_spot_number=min_spot_number, shape=image.shape)
-        else : new_clusters = None
+    new_spots = np.array(Viewer.layers['single spots'].data, dtype= int)
 
-    except Exception as error :
-        new_spots = spots
-        new_clusters = clusters
-        raise error
+    if type(clusters) != type(None) :
+        if len(clusters) > 0 : 
+            new_clusters = np.array(Viewer.layers['foci'].data, dtype= int)
+            new_clusters = _update_clusters(new_clusters, new_spots, voxel_size=voxel_size, cluster_size=cluster_size, min_spot_number=min_spot_number, shape=image.shape)
+    else : new_clusters = None
 
     return new_spots, new_clusters
 
@@ -168,25 +138,27 @@ def show_segmentation(
     Viewer = napari.Viewer(ndisplay=2, title= 'Show segmentation', axis_labels=['z','y','x'] if dim == 3 else ['y', 'x'], show= False)
     
     # Adding channels
-    Viewer.add_image(nuc_image, name= "nucleus signal", blending= 'additive', colormap='blue', contrast_limits=[nuc_image.min(), nuc_image.max()])
-    Viewer.add_labels(nuc_label, opacity= 0.5, blending= 'additive')
+    nuc_signal_layer = Viewer.add_image(nuc_image, name= "nucleus signal", blending= 'additive', colormap='blue', contrast_limits=[nuc_image.min(), nuc_image.max()])
+    nuc_label_layer = Viewer.add_labels(nuc_label, opacity= 0.5, blending= 'additive', name= 'nucleus_label',)
+    nuc_label_layer.preserve_labels = True
     
     #Adding labels
     if type(cyto_image) != type(None) : Viewer.add_image(cyto_image, name= "cytoplasm signal", blending= 'additive', colormap='red', contrast_limits=[cyto_image.min(), cyto_image.max()])
-    if type(cyto_label) != type(None) and not np.array_equal(cyto_label, nuc_label): Viewer.add_labels(cyto_label, opacity= 0.4, blending= 'additive')
+    if (type(cyto_label) != type(None) and not np.array_equal(cyto_label, nuc_label) ) or (type(cyto_label) != type(None) and cyto_label.max() == 0): 
+        cyto_label_layer = Viewer.add_labels(cyto_label, opacity= 0.4, blending= 'additive', name= 'cytoplasm_label')
+        cyto_label_layer.preserve_labels = True
     
     #Launch Napari
     Viewer.show(block=False)
     napari.run()
 
-    new_nuc_label = Viewer.layers[1].data
-    if type(cyto_label) != type(None) and not np.array_equal(cyto_label, nuc_label) : new_cyto_label = Viewer.layers[3].data
+
+    new_nuc_label = Viewer.layers['nucleus_label'].data
+    if type(cyto_label) != type(None) and not np.array_equal(cyto_label, nuc_label) : new_cyto_label = Viewer.layers['cytoplasm_label'].data
     else : new_cyto_label = new_nuc_label
 
     return new_nuc_label, new_cyto_label
 
-
-
 def threshold_selection(
         image : np.ndarray,
         filtered_image : np.ndarray,
@@ -195,7 +167,7 @@ def threshold_selection(
         ) :
     
     """
-    To view code for spot selection please have a look at magicgui instance created with `detection._create_threshold_slider` which is then passed to this napari wrapper as 'threshold_slider' argument.
+    To view code for spot selection have a look at magicgui instance created with `detection._create_threshold_slider` which is then passed to this napari wrapper as 'threshold_slider' argument.
     """
     
     Viewer = napari.Viewer(title= "Small fish - Threshold selector", ndisplay=2, show=True)
@@ -218,12 +190,13 @@ def threshold_selection(
 
     Viewer.window.add_dock_widget(threshold_slider, name='threshold_selector')
     threshold_slider() #First occurence with auto or entered threshold.
+    
     napari.run()
 
-    spots = Viewer.layers[-1].data.astype(int)
+    spots = Viewer.layers['single spots'].data.astype(int)
     if len(spots) == 0 :
-        threshold = filtered_image.max()
+        pass
     else :
-        threshold = Viewer.layers[-1].properties.get('threshold')[0]
+        threshold = Viewer.layers['single spots'].properties.get('threshold')[0]
 
     return spots, threshold