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@ -1,207 +1,315 @@ |
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from pims import Frame |
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from pims.base_frames import FramesSequenceND |
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# -*- coding: utf-8 -*- |
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import struct |
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from nd2reader.exceptions import EmptyFileError, InvalidFileType |
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from nd2reader.parser import Parser |
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import array |
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import six |
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import warnings |
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from pims.base_frames import Frame |
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import numpy as np |
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import numpy as np |
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from nd2reader.common import get_version, read_chunk |
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from nd2reader.exceptions import InvalidVersionError |
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from nd2reader.label_map import LabelMap |
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from nd2reader.raw_metadata import RawMetadata |
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class ND2Reader(FramesSequenceND): |
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"""PIMS wrapper for the ND2 parser. |
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This is the main class: use this to process your .nd2 files. |
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""" |
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class_priority = 12 |
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class Parser(object): |
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"""Parses ND2 files and creates a Metadata and driver object. |
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def __init__(self, filename): |
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super(ND2Reader, self).__init__() |
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""" |
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CHUNK_HEADER = 0xabeceda |
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CHUNK_MAP_START = six.b("ND2 FILEMAP SIGNATURE NAME 0001!") |
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CHUNK_MAP_END = six.b("ND2 CHUNK MAP SIGNATURE 0000001!") |
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if not filename.endswith(".nd2"): |
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raise InvalidFileType("The file %s you want to read with nd2reader does not have extension .nd2." % filename) |
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supported_file_versions = {(3, None): True} |
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self.filename = filename |
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def __init__(self, fh): |
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self._fh = fh |
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self._label_map = None |
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self._raw_metadata = None |
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self.metadata = None |
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# first use the parser to parse the file |
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self._fh = open(filename, "rb") |
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self._parser = Parser(self._fh) |
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# First check the file version |
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self.supported = self._check_version_supported() |
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# Setup metadata |
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self.metadata = self._parser.metadata |
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# Parse the metadata |
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self._parse_metadata() |
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# Set data type |
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self._dtype = self._parser.get_dtype_from_metadata() |
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def calculate_image_properties(self, index): |
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"""Calculate FOV, channels and z_levels |
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# Setup the axes |
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self._setup_axes() |
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Args: |
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index(int): the index (which is simply the order in which the image was acquired) |
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# Other properties |
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self._timesteps = None |
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Returns: |
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tuple: tuple of the field of view, the channel and the z level |
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@classmethod |
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def class_exts(cls): |
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"""Let PIMS open function use this reader for opening .nd2 files |
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""" |
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field_of_view = self._calculate_field_of_view(index) |
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channel = self._calculate_channel(index) |
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z_level = self._calculate_z_level(index) |
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return field_of_view, channel, z_level |
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def get_image(self, index): |
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""" |
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""" |
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return {'nd2'} | super(ND2Reader, cls).class_exts() |
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Creates an Image object and adds its metadata, based on the index (which is simply the order in which the image |
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was acquired). May return None if the ND2 contains multiple channels and not all were taken in each cycle (for |
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example, if you take bright field images every minute, and GFP images every five minutes, there will be some |
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indexes that do not contain an image. The reason for this is complicated, but suffice it to say that we hope to |
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eliminate this possibility in future releases. For now, you'll need to check if your image is None if you're |
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doing anything out of the ordinary. |
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def close(self): |
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"""Correctly close the file handle |
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Args: |
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index(int): the index (which is simply the order in which the image was acquired) |
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""" |
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if self._fh is not None: |
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self._fh.close() |
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Returns: |
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Frame: the image |
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def _get_default(self, coord): |
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""" |
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field_of_view, channel, z_level = self.calculate_image_properties(index) |
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channel_offset = index % len(self.metadata["channels"]) |
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image_group_number = int(index / len(self.metadata["channels"])) |
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frame_number = self._calculate_frame_number(image_group_number, field_of_view, z_level) |
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try: |
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try: |
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return self.default_coords[coord] |
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except KeyError: |
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return 0 |
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timestamp, image = self._get_raw_image_data(image_group_number, channel_offset, self.metadata["height"], |
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self.metadata["width"]) |
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except (TypeError): |
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return Frame([], frame_no=frame_number, metadata=self._get_frame_metadata()) |
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else: |
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return Frame(image, frame_no=frame_number, metadata=self._get_frame_metadata()) |
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def get_image_by_attributes(self, frame_number, field_of_view, channel, z_level, height, width): |
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"""Gets an image based on its attributes alone |
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def get_frame_2D(self, c=0, t=0, z=0, x=0, y=0, v=0): |
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"""Gets a given frame using the parser |
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Args: |
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Args: |
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x: The x-index (pims expects this) |
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y: The y-index (pims expects this) |
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c: The color channel number |
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t: The frame number |
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z: The z stack number |
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v: The field of view index |
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frame_number: the frame number |
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field_of_view: the field of view |
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channel_name: the color channel name |
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z_level: the z level |
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height: the height of the image |
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width: the width of the image |
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Returns: |
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Returns: |
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pims.Frame: The requested frame |
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Frame: the requested image |
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""" |
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""" |
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# This needs to be set to width/height to return an image |
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x = self.metadata["width"] |
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y = self.metadata["height"] |
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frame_number = 0 if frame_number is None else frame_number |
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field_of_view = 0 if field_of_view is None else field_of_view |
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channel = 0 if channel is None else channel |
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z_level = 0 if z_level is None else z_level |
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return self._parser.get_image_by_attributes(t, v, c, z, y, x) |
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image_group_number = self._calculate_image_group_number(frame_number, field_of_view, z_level) |
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try: |
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timestamp, raw_image_data = self._get_raw_image_data(image_group_number, channel, |
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height, width) |
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except (TypeError): |
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return Frame([], frame_no=frame_number, metadata=self._get_frame_metadata()) |
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else: |
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return Frame(raw_image_data, frame_no=frame_number, metadata=self._get_frame_metadata()) |
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@staticmethod |
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def get_dtype_from_metadata(): |
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"""Determine the data type from the metadata. |
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For now, always use float64 to prevent unexpected overflow errors when manipulating the data (calculating sums/ |
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means/etc.) |
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@property |
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def parser(self): |
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""" |
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Returns the parser object. |
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Returns: |
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Parser: the parser object |
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""" |
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""" |
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return self._parser |
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return np.float64 |
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@property |
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def pixel_type(self): |
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"""Return the pixel data type |
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def _check_version_supported(self): |
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"""Checks if the ND2 file version is supported by this reader. |
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Returns: |
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Returns: |
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dtype: the pixel data type |
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bool: True on supported |
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""" |
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""" |
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return self._dtype |
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major_version, minor_version = get_version(self._fh) |
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supported = self.supported_file_versions.get( |
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(major_version, minor_version)) or self.supported_file_versions.get((major_version, None)) |
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@property |
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def timesteps(self): |
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"""Get the timesteps of the experiment |
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if not supported: |
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print("Warning: No parser is available for your current ND2 version (%d.%d). " % ( |
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major_version, minor_version) + "This might lead to unexpected behaviour.") |
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Returns: |
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np.ndarray: an array of times in milliseconds. |
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return supported |
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def _parse_metadata(self): |
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"""Reads all metadata and instantiates the Metadata object. |
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""" |
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""" |
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if self._timesteps is None: |
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return self.get_timesteps() |
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return self._timesteps |
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# Retrieve raw metadata from the label mapping |
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self._label_map = self._build_label_map() |
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self._raw_metadata = RawMetadata(self._fh, self._label_map) |
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self.metadata = self._raw_metadata.__dict__ |
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self.acquisition_times = self._raw_metadata.acquisition_times |
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@property |
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def events(self): |
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"""Get the events of the experiment |
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def _build_label_map(self): |
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""" |
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Every label ends with an exclamation point, however, we can't directly search for those to find all the labels |
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as some of the bytes contain the value 33, which is the ASCII code for "!". So we iteratively find each label, |
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grab the subsequent data (always 16 bytes long), advance to the next label and repeat. |
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Returns: |
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Returns: |
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iterator of events as dict |
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LabelMap: the computed label map |
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""" |
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""" |
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# go 8 bytes back from file end |
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self._fh.seek(-8, 2) |
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chunk_map_start_location = struct.unpack("Q", self._fh.read(8))[0] |
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self._fh.seek(chunk_map_start_location) |
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raw_text = self._fh.read(-1) |
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return LabelMap(raw_text) |
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return self._get_metadata_property("events") |
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def _calculate_field_of_view(self, index): |
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"""Determines what field of view was being imaged for a given image. |
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Args: |
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index(int): the index (which is simply the order in which the image was acquired) |
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@property |
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def frame_rate(self): |
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"""The (average) frame rate |
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Returns: |
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Returns: |
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float: the (average) frame rate in frames per second |
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int: the field of view |
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""" |
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""" |
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total_duration = 0.0 |
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images_per_cycle = len(self.metadata["z_levels"]) * len(self.metadata["channels"]) |
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return int((index - (index % images_per_cycle)) / images_per_cycle) % len(self.metadata["fields_of_view"]) |
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for loop in self.metadata['experiment']['loops']: |
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total_duration += loop['duration'] |
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def _calculate_channel(self, index): |
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"""Determines what channel a particular image is. |
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if total_duration == 0: |
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total_duration = self.timesteps[-1] |
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Args: |
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index(int): the index (which is simply the order in which the image was acquired) |
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if total_duration == 0: |
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raise ValueError('Total measurement duration could not be determined from loops') |
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Returns: |
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string: the name of the color channel |
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return self.metadata['num_frames'] / (total_duration/1000.0) |
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""" |
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return self.metadata["channels"][index % len(self.metadata["channels"])] |
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def _get_metadata_property(self, key, default=None): |
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if self.metadata is None: |
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return default |
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def _calculate_z_level(self, index): |
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"""Determines the plane in the z-axis a given image was taken in. |
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if key not in self.metadata: |
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return default |
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In the future, this will be replaced with the actual offset in micrometers. |
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if self.metadata[key] is None: |
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return default |
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Args: |
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index(int): the index (which is simply the order in which the image was acquired) |
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return self.metadata[key] |
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Returns: |
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The z level |
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def _setup_axes(self): |
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"""Setup the xyctz axes, iterate over t axis by default |
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""" |
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return self.metadata["z_levels"][int( |
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((index - (index % len(self.metadata["channels"]))) / len(self.metadata["channels"])) % len( |
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self.metadata["z_levels"]))] |
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def _calculate_image_group_number(self, frame_number, fov, z_level): |
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""" |
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""" |
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self._init_axis_if_exists('x', self._get_metadata_property("width", default=0)) |
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self._init_axis_if_exists('y', self._get_metadata_property("height", default=0)) |
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self._init_axis_if_exists('c', len(self._get_metadata_property("channels", default=[])), min_size=2) |
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self._init_axis_if_exists('t', len(self._get_metadata_property("frames", default=[]))) |
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self._init_axis_if_exists('z', len(self._get_metadata_property("z_levels", default=[])), min_size=2) |
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self._init_axis_if_exists('v', len(self._get_metadata_property("fields_of_view", default=[])), min_size=2) |
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Images are grouped together if they share the same time index, field of view, and z-level. |
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if len(self.sizes) == 0: |
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raise EmptyFileError("No axes were found for this .nd2 file.") |
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Args: |
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frame_number: the time index |
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fov: the field of view number |
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z_level: the z level number |
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# provide the default |
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self.iter_axes = self._guess_default_iter_axis() |
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Returns: |
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int: the image group number |
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self._register_get_frame(self.get_frame_2D, 'yx') |
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""" |
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z_length = len(self.metadata['z_levels']) |
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z_length = z_length if z_length > 0 else 1 |
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fields_of_view = len(self.metadata["fields_of_view"]) |
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fields_of_view = fields_of_view if fields_of_view > 0 else 1 |
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def _init_axis_if_exists(self, axis, size, min_size=1): |
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if size >= min_size: |
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self._init_axis(axis, size) |
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return frame_number * fields_of_view * z_length + (fov * z_length + z_level) |
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def _guess_default_iter_axis(self): |
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def _calculate_frame_number(self, image_group_number, field_of_view, z_level): |
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""" |
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""" |
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Guesses the default axis to iterate over based on axis sizes. |
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Images are in the same frame if they share the same group number and field of view and are taken sequentially. |
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Args: |
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image_group_number: the image group number (see _calculate_image_group_number) |
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field_of_view: the field of view number |
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z_level: the z level number |
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Returns: |
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Returns: |
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the axis to iterate over |
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""" |
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""" |
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priority = ['t', 'z', 'c', 'v'] |
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found_axes = [] |
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for axis in priority: |
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try: |
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current_size = self.sizes[axis] |
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except KeyError: |
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continue |
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return (image_group_number - (field_of_view * len(self.metadata["z_levels"]) + z_level)) / ( |
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len(self.metadata["fields_of_view"]) * len(self.metadata["z_levels"])) |
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if current_size > 1: |
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return axis |
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@property |
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def _channel_offset(self): |
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""" |
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Image data is interleaved for each image set. That is, if there are four images in a set, the first image |
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will consist of pixels 1, 5, 9, etc, the second will be pixels 2, 6, 10, and so forth. |
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found_axes.append(axis) |
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Returns: |
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|
dict: the channel offset for each channel |
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return found_axes[0] |
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""" |
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|
return {channel: n for n, channel in enumerate(self.metadata["channels"])} |
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def _remove_unwanted_bytes(self, image_group_data, image_data_start, height, width): |
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|
# Remove unwanted 0-bytes that can appear in stitched images |
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|
number_of_true_channels = int(len(image_group_data[4:]) / (height * width)) |
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|
unwanted_bytes_len = (len(image_group_data[image_data_start:]))%(height*width) |
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|
if unwanted_bytes_len: |
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|
warnings.warn('Identified unwanted bytes in the ND2 file, possibly stitched.') |
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|
byte_ids = range(image_data_start+height*number_of_true_channels, len(image_group_data)-unwanted_bytes_len+1, height*number_of_true_channels) |
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|
if all([0 == image_group_data[byte_ids[i]+i] for i in range(len(byte_ids))]): |
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|
warnings.warn('All unwanted bytes are zero-bytes, correctly removed.') |
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|
|
for i in range(len(byte_ids)): |
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|
del image_group_data[byte_ids[i]] |
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|
def _get_raw_image_data(self, image_group_number, channel_offset, height, width): |
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|
|
"""Reads the raw bytes and the timestamp of an image. |
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|
|
def get_timesteps(self): |
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|
|
"""Get the timesteps of the experiment |
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|
|
Args: |
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|
|
image_group_number: the image group number (see _calculate_image_group_number) |
|
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|
|
channel_offset: the number of the color channel |
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|
|
height: the height of the image |
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|
|
width: the width of the image |
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|
|
Returns: |
|
|
Returns: |
|
|
np.ndarray: an array of times in milliseconds. |
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|
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|
|
|
""" |
|
|
""" |
|
|
if self._timesteps is not None and len(self._timesteps) > 0: |
|
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|
|
|
return self._timesteps |
|
|
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|
|
|
|
|
chunk = self._label_map.get_image_data_location(image_group_number) |
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|
|
data = read_chunk(self._fh, chunk) |
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|
|
|
# All images in the same image group share the same timestamp! So if you have complicated image data, |
|
|
|
|
|
# your timestamps may not be entirely accurate. Practically speaking though, they'll only be off by a few |
|
|
|
|
|
# seconds unless you're doing something super weird. |
|
|
|
|
|
timestamp = struct.unpack("d", data[:8])[0] |
|
|
|
|
|
image_group_data = array.array("H", data) |
|
|
|
|
|
image_data_start = 4 + channel_offset |
|
|
|
|
|
|
|
|
|
|
|
# The images for the various channels are interleaved within the same array. For example, the second image |
|
|
|
|
|
# of a four image group will be composed of bytes 2, 6, 10, etc. If you understand why someone would design |
|
|
|
|
|
# a data structure that way, please send the author of this library a message. |
|
|
|
|
|
number_of_true_channels = int(len(image_group_data[4:]) / (height * width)) |
|
|
|
|
|
self._remove_unwanted_bytes(image_group_data, image_data_start, height, width) |
|
|
|
|
|
try: |
|
|
|
|
|
image_data = np.reshape(image_group_data[image_data_start::number_of_true_channels], (height, width)) |
|
|
|
|
|
except ValueError: |
|
|
|
|
|
image_data = np.reshape(image_group_data[image_data_start::number_of_true_channels], (height, int(round(len(image_group_data[image_data_start::number_of_true_channels])/height)))) |
|
|
|
|
|
|
|
|
|
|
|
# Skip images that are all zeros! This is important, since NIS Elements creates blank "gap" images if you |
|
|
|
|
|
# don't have the same number of images each cycle. We discovered this because we only took GFP images every |
|
|
|
|
|
# other cycle to reduce phototoxicity, but NIS Elements still allocated memory as if we were going to take |
|
|
|
|
|
# them every cycle. |
|
|
|
|
|
if np.any(image_data): |
|
|
|
|
|
return timestamp, image_data |
|
|
|
|
|
|
|
|
|
|
|
# If a blank "gap" image is encountered, generate an array of corresponding height and width to avoid |
|
|
|
|
|
# errors with ND2-files with missing frames. Array is filled with nan to reflect that data is missing. |
|
|
|
|
|
else: |
|
|
|
|
|
empty_frame = np.full((height, width), np.nan) |
|
|
|
|
|
warnings.warn('ND2 file contains gap frames which are represented by np.nan-filled arrays; to convert to zeros use e.g. np.nan_to_num(array)') |
|
|
|
|
|
return timestamp, image_data |
|
|
|
|
|
|
|
|
|
|
|
def _get_frame_metadata(self): |
|
|
|
|
|
"""Get the metadata for one frame |
|
|
|
|
|
|
|
|
self._timesteps = np.array(list(self._parser._raw_metadata.acquisition_times), dtype=np.float) * 1000.0 |
|
|
|
|
|
|
|
|
Returns: |
|
|
|
|
|
dict: a dictionary containing the parsed metadata |
|
|
|
|
|
|
|
|
return self._timesteps |
|
|
|
|
|
|
|
|
""" |
|
|
|
|
|
return self.metadata |