Data Interpretation

Data Interpretation

The output format varies depending on how the acquisition is performed:

Normal Acquisition

Normal acquisition refers to taking images without using any advanced features (e.g. light program). Normally each normal acquisition generates 8 files:
  1. .locb: a binary representation of the localization data.
  2. .nimb: propriatary instrument data to ONI.
  3. .tif: the raw images, either the full camera chip or a stitch of the two channels.
  4. acq.nim: the acquisition metafile.
  5. camcal.nim: the camera metafile.
  6. map.nim: the channel mapping data.
  7. thumbnail.png: a thumbnail of the localization data.
  8. locImage.tif: a rendering of the localization data. This file will be generated if the option Save A Rendered Image Of Localizations is checked in Settings > User Settings.
The result from a normal acquisition contains two channels when read in NimOS:
  1. Channel 0/Channel 1: the raw data (i.e. the wide-field images), in which Channel 0 corresponds to the left channel and Channel 1 corresponds to the right channel.
  2. G0 - Left/G0 - Right: the localization data, in which G0 - Left corresponds to the left channel and G0 - Right corresponds to the right channel.
For example, for Alexa 647, you should expect to see the raw data in Channel 1 and the localization data in G0 - Right.

Light Program Acquisition

Light program acquisition refers to taking images using the light program. Similar to a normal acquisition, normally each light program acquisition generates 8 files.
The result from a light program acquisition contains several channels depending on the program when read in NimOS:
For example, for standard ONI 3-color EV imaging (CF 488A, CF 568, and CF 647) with a 640-split system, you should expect to see:
  1. The raw data of CF 647 in Channel 1 and the corresponding localization data in G0 - Right.
  2. The raw data of CF 568 in Channel 2 and the corresponding localization data in G0 - Left.
  3. The raw data of CF 488A in Channel 4 and the corresponding localization data in G1 - Left.
The interpretation differs for a 560-split system and for detailed instructions, check here.

Multi-Acquisition

Multi-acquisition refers to taking images using the multi-acquisition program. Similar to a normal acquisition, normally each multi-acquisition generates 8 files per laser/position/timepoint. If Combine Datasets is checked, additional files will be generated, in which the two channels are combined into a Hyperstack.
The result from a multi-acquisition contains two channels per laser/position/timepoint when read in NimOS.

Confocal Acquisition

Confocal acquisition refers to taking images with confocal enabled. Besides the 7 files generated from a normal acquisition (without the ".locb" file), normally each confocal acquisition generates additional 5 files:

  1. confocal.tif: the confocal image of each channel.
  2. widefield.tif: the wide-field image of each channel.
  3. scparams.nim: the confocal processing data.
Similar to a normal acquisition, the result from a confocal acquisition contains two channels when read in NimOS, however, both G0 - Left and G0 - Right are "empty". Click on Run Localization to run localization.

3D Acquisition

3D acquisition refers to taking images with the 3D lens engaged. Similar to a normal acquisition, normally each 3D acquisition generates 8 files, in which the Z positions are encoded in the ".locb" file.
Similar to a normal acquisition, the result from a 3D acquisition contains two channels when read in NimOS. However, the filter function will contain filtering options for the Z axis:
If at any point you are having issues with the interpretation process, please do not hesitate to contact the CX team.

    • Related Articles

    • Data Metadata

      Two ways to read the metadata of the current data: Either click on Data > View Acquisition Data to show the metafile: Or open the corresponding "acq.nim" file to show the metadata:

    • Data Import

      Instructions: In Analyze, click on Load Data. Navigate to the folder of interest and select either the Tiff file or the Locb File to import. Discussion: If the dataset is modified via 3rd party software such as ImageJ, it may not be compatible with ...

    • Data Export

      There are different ways to save/export data from NimOS: Copy To Clipboard (Raw) Instructions: Right-click on any of the channels and click on Copy To Clipboard: Paste it into any other data viewer (e.g. ImageJ, Paint, Excel...) to visualize and save ...

    • NimOS Analysis

      NimOS provides functions for data analysis: Analysis: Tools Analysis: Viewing Options Analysis: Filter Analysis: Localizations Analysis: Drift Correction

    • Analysis: Drift Correction

      A dSTORM acquisition is usually long and subject to drift (the image being slowly shifted to one direction) caused by, for example, temperature fluctuation. Drift Correction allows correcting for the drift as post-processing. There are two methods to ...