The newly redesigned SlideBook™ 4.2 application comes complete with all of the necessary tools for imaging application ranging from simple 2D and timelapse imaging to multichannel 3D, multipoint, and 4D imaging. SlideBook™ features the Data Slide architecture, which allows the user to store related images together and simplifies both data analysis and archive retrieval. SlideBook's unique software architecture is capable of 2D, 3D, 4D and 6D image acquisition. Morphometrics, time-lapse and an advanced statistics engine are included as standard. Optional modules are available for deconvolution, ratiometric-fluorimetry (FRET, pH, calcium), and TTL synchronization.The data structure in SlideBook also facilitates the accurate recording of image characteristics by automatically storing all of the information from image capture along with the individual images.
Features and Benefits
- For use with Olympus BX, BX2, IX, IX2 and (DSU) Spinning Disk Confocal microscopes.
- Controls all hardware types associated with digital microscopy to enable fully-automated scope control including fluorescence shutters, filter wheels, rapid wavelength changers, X-Y stages, piezo-controlled Z-Axis systems and focus motors.
- Native 3D image software architecture allows user-friendly viewing, manipulation and analysis of multi-dimensional and multi-channel data without the need for scripts or macros.
- 2D, 3D, 4D and 6D data capture, visualization, segmentation (mask and sub-mask creation), and analysis tools.
- Full camera control for most scientific grade CCD cameras. Supports advanced camera functions such as on-line control of digitizer speed, gain, offset, intensification, sub-region readout and binning. Simultaneous camera control.
- Unique file system allows users to store related images as slides for rapid analysis and archive retrieval. Imaging parameters are automatically recorded and permanently stored with image data.
- Morphometrics including intensity segmentation, tools for local segmentation, volume, surface area, 3D distance (linear and non-linear).
- No neighbors deconvolution, common filters, photobleach correction, flat field correction and custom convolution kernels.
- Control and live feedback during image acquisition including:
- graphing of wavelength intensities
- calibrated ion concentrations for multiple ROIs
- capture pause
- interactive refocusing
- ROI adjustment
- event marking
- Advanced statistics engine for data quantification
- Montage creation
- Multi-point 2D, 3D, 4D and 6D
- Supported imaging techniques include:
- Bright Field
- Dark Field
- Phase Contrast
- DIC (differential interference contrast)
- TIRF (total internal reflection microscopy)
- IRM (internal reflection microscopy)
The 3D Deconvolution module adds a 3D deblurring and an image restoration algorithm to SlideBook. The nearest neighbor and constrained iterative deconvolution algorithms have been optimized for speed and performance. The module includes an interactive guide for measuring point spread functions (PSF) and a PSF database. The database can store multiple PSFs for each configuration, allowing easy selection of the appropriate PSF.
Nearest Neighbors Deconvolution
Our Nearest Neighbors deconvolution is a rapid way to deblur fluorescence data. The algorithm uses the plane above and below the plane of interest to compute and subtract the fraction of the data that is out-of-focus information.
Constrained Iterative Deconvolution
Our Constrained Iterative (CI) deconvolution is a true image restoration tool. Based on the algorithm developed by David Agard at UCSF, our CI deconvolution can quantitatively reassign out-of-focus information in 3-D data while improving both axial and lateral data resolution.
The FRD module for SlideBook™ 4.0 is the first commercially available software specifically designed for FRET analysis. This revolutionary software was developed in collaboration with a number of leading biologists whose laboratories focus on FRET. The FRET module contains analysis tools for both sensitized emission FRET and acceptor photobleaching FRET. These tools include:
- Channel Bleedthrough Calculation
- Background Subtraction
- Corrected FRET Image Generation (Herman Equation)
- Single Pixel to Whole Object Energy Transfer Measurement
- Support for 2-D, 3-D and 4-D Data
The module can be combined with image capture and other on-line modules for real-time FRET analysis. In addition, SlideBook™ supports dual camera capture and the Multi-Spec Microimager device from Optical Insights for true real-time FRET image acquisition. The measurement of free ion concentrations in living cells provides insight into cellular processes such as activation and compartmentalization. Ratiometric ion indicators change their excitation or emission profiles when bound to specific ions. These indicators allow the real-time measurement of ion concentrations in living cells. :This module supports a wide array of ion indicators including FURA-2, Indo-1 and BCECF. The module includes a variety of specific features including:
- Real-time ratio computation
- Real-time graphing of ratios and/or raw channels
- Real-time tracking of up to 20 ROI
- Simultaneous imaging of two ratio indicators
- Calibration guide for computing nM calcium concentration by pixel
- Analytical Graphing tools
Particle Tracking Module
The evolution of digital microscopy has placed increasing emphasis on the quantitative information contained within collected images. The Statistics Engine for SlideBook™ has been completely reworked for version 4.0 to provide researchers with a complete suite of analytical tools specifically designed for quantifying data from microscopes. Many new statistics, including cross-channel colocalization and skeleton-based features. The new Statistics Engine includes:
- Plug-in architecture for end-user writable features
- Automatic methods for generating and storing statistics to aid in gathering large amounts of data
- Advanced clustering techniques to help define populations of objects and images
- Many new statistics, including cross-channel colocalization and skeleton-based features
The all new 4D analysis and particle tracking module for SlideBook 4.0 contains powerful tools tracking individual 3D objects in 4-D datasets. The particle tracking engine is based on individual object paths. Individual objects will have a unique path through time and can therefore be followed.The particle tracking module can also correlate objects with respect to one another and generate statistics about each object at each time-point.
Object paths can be traced either automatically using computational algorithms or through manual delineation. The particle tracking module also offers complete flexibility for combining both automated and manual tools to optimize path detection for any given sample. Once object paths have been traced, there are a variety of visualization tools to help explore the objects. In the OpenGL-based 3DpathView, for example, an individual path can be selected and individually rendered over time. This rendering can be rotated freely and played as a movie (in either direction) or in single frame increments. As users explore the data, they can manipulate the path, either to prune a split/merge as objects move, or get statistics for a given object.
The statistics generated by the particle tracker include everything from velocity and acceleration of objects to a description of randomness of an object's motion. These statistics can be automatically generated for all objects/paths or statistics can be generated on a path-by-path basis. In addition, all delineations, processes and analyses in the particle tracker are automatable. Each step in the tracking scheme can be stored and retrieved from a methods database, so that a series of commands can be easily written and retrieved for use in future experiments.
Multi-Well Plate Module
SlideBook™ 4.0 features a new, easy-to-use lpate reading module. This module enables SlideBook™ to capture data from a wide range of multi-well plates, including 24, 96, and 384-well plates.
This module gives the researcher complete control over all imaging parameters. An experiment can include any combination of SlideBook™ supported imaging modes, including multi-spectral fluorescence and combination fluorescence/transmitted light imaging. In addition, this module includes an optimized auto focus algorithm to ensure high quality image data. The researcher can select the range of wells to be images, along with the location and number of images to be taken per well. These parameters can be saved as a template, making it fast and easy to image multiple plates under identical conditions.
The rapid collection of 4-D data requires the precise timing and synchronization of all hardware involved in image capture. The TTL module for SlideBook™ 4.0 is a combination of hardware and software specifically designed to optimize the recording of 4-D data and synchronizing external devices such as perfusion systems and electrophysiology equipment.
Normally, hardware device control relies on serial connections between the computer and the specific piece of hardware. While serial control can accurately control single devices, it is difficult to simultaneously coordinate multiple devices through this interface. The Rapid 4D Synchronization module utilizes analog pulses of square waves (TTL pulses) to simultaneously trigger multiple devices for operation. The module comes complete with all of the necessary hardware for controlling devices through TTL pulses including multiple analog in and out connectors, PCI board and cables.