General Information

Deep Thought is a Windows 7 64-bit image Dell Precision T7600 with dual Intel XEON CPU E5-2620 2 GHz processors (6 cores 12 logical partitions), 160 GB RAM and an AMD Radeon HD 7800 2 GB VRAM graphics card. This computer has almost the maximum amount of RAM possible in Windows 7, which makes it very suitable for memory intensive tasks like image deconvolution. The deconvolution software package Huygens Essential is installed on this machine to make best use of the RAM. There is also a full copy of Carl Zeiss's Zen software installed (both Black and Blue editions) with a module for processing Airyscan data offline and a copy of Nikon NIS-Elements with an offline package for analysing NSTORM molecular localisation data.

Software on Deep Thought

Huygens Essential

Deconvolution software. Deep Thought is currently running version 16.05 (built on October 2016) with licences for deconvolving data from widefield, confocal, spinning disk and multiphoton microscopes. The software also has the PSF distiller, which allows you to create point spread functions from sub-resolution bead specimens imaged on your own microscope. Note that the Orac analysis computer has a Huygens sub-licence linked to the STED microscope, which can deconvolve STED and confocal data but not the other modalities listed above. For widefield, spinning disk and multiphoton you will need to use Huygens on Deep Thought.

 Microsoft Remote Desktop compatible

Carl Zeiss Zen

Deep Thought is currently running Zen 2.1 Sp3 version 14.0.0.201. The copy on this machine is an offline licence linked to the Multiphoton. It has analysis modules relevant to that machine. The most important being probably: FRAP analysis, FCS analysis tools (FCS Basis, FCS Extended, FCS PCH) and Airyscan processing. It is also able to perform linear unmixing of spectral imaging data from Zeiss confocal microscopes.

If you need to process structured illumination imaging (SIM) data from the Elyra microscope the modules for that are only on the Zen analysis computer; not on Deep Thought.

 Microsoft Remote Desktop incompatible. If you launch Zen through RDP then it will open only the Lite version and the advanced analysis tools will not be available.

Nikon NIS-Elements 5.01

This offline licence is associated with the Nikon time-lapses and has the standard set of analysis tools but its main advantage is an offline licence for analysis of NSTORM data.

 Microsoft Remote Desktop incompatible

SlideBook 6.0.15

Offline software for 3i spinning disc confocal microscope.

 Microsoft Remote Desktop compatible

Volocity 6.3

Key features: 3D surface rendering, volume rendering and ray tracing. Identify and measure 3D objects and object properties. Deconvolution.

Volocity natively analyses data sets in 3D rather than as stacks of 2D slices. This means that measurements automatically refer to 3D objects rather than single slices and are given in 3D units (e.g. cubic microns). Volocity consists of a core programs that allows you to open your data, carry out some basic processing and export in a variety of formats. If you want to carry out advanced processing in Volocity you need to use one of the analysis modules: Restoration, Visualization and Quantitation. Restoration is a deconvolution package of most use with high quality confocal data. Visualization is a 3D reconstruction and rendering package with volume and surface rendering tools and a ray tracer. Quantitation is a measurement package that can be used to apply a 'pipeline' of analysis steps to process data and extract measurements. Basically this means thresholding and segmentation, binary style procedures like erosion and dilation, object identification, filtering by object features, measurement and colocalisation, etc.

I find that the Restoration module is not great at deconvolving data with lower signal to noise ratios, so I tend to use Huygens and AutoQuant for deconvolution. It is difficult to set up an analysis pipeline using the Quantitation but all such tools are difficult and require a lot of trial and error, so this isn't unusual. The analysis and charts tools in the Quantitation module are not intuitive at all so most of the time I export data as a spreadsheet for analysis in other packages (e.g. Excel). The Tracking tools for measuring objects moving over time seem to be a lot more flexible and user friendly in Imaris.

 Microsoft Remote Desktop compatible

Free analysis software and browsers

Fiji (ImageJ)

The Fiji installed on this computer currently has a number of locally installed modules in addition to the standard Fiji plugins that are automatically updated. Here are a few of the plugins available.

• The NanoJ toolset including NanoJ-SQUIRREL and NanoJ-SRRF
• SIMCheck, for validating data from structured illumination microscopes
• ThunderSTORM, for analysis of STORM data
• The ImageScience toolset, including NeuronJ
• HDF5 for opening HDF5 files, which is a file format for storing and viewing very large images (like tiled slides)

You can also install these plugins for your own copy of Fiji, although you might not be able to run all of them efficiently if you don't have a powerful enough computer. See the following link for details of the range of update sites that can be used to update different plugins in Fiji, but bear in mind the caveat at the top of the linked page warning you about the risk of breaking your Fiji installation.

 Microsoft Remote Desktop compatible

Leica LAS AF Lite and LAS X

LAS AF Lite version 2.6.3. LAS X version 3.0.1.15878. Free browser versions of Leica's confocal microscope software have some basic processing and analysis capabilities but are probably most useful for browsing images and stacks prior to deconvolution or further processing in other software. Data from the STED will only open properly in the more recent LAS X software. LIF files from the older confocal microscopes will open in both LAS AF Lite and LAS X, but the former is probably preferable, since re-saving the data in LAS X will make the file incompatible with the machine it was acquired on.

 Microsoft Remote Desktop compatible

FV10-ASW Viewer

Free browser version for OIF and OIB files from the Olympus Fluoview FV1200 confocal microscope.

 Microsoft Remote Desktop compatible