IVIS Spectrum Tips & Tricks
Tips / Shortcuts
Settings Cheat Sheet for the IVIS Spectrum
The F number refers to the aperture, or size of the hole that lets light fall onto the CCD in the camera. F1 is WIDE OPEN, whereas F8 is small.
What IS binning? Pixel binning is a clocking scheme used to combine the charge collected by several adjacent CCD pixels, and is designed to reduce noise and improve the signal-to-noise ratio and frame rate of digital cameras. The IVIS has 3 different Binning settings – Low, Medium & High.
In the context of image processing, binning is the procedure of combining a cluster of into a single pixel. As such, in 2×2 binning, an array of 4 pixels becomes a single larger pixel, reducing the overall number of pixels.
This aggregation, reducing the number of data (with a loss of information), facilitates the analysis. For instance, binning the data may also reduce the impact of read noise on the processed image (at the cost of a lower resolution).
Thus, the HIGHER the binning, the LOWER the resolution and the QUICKER the analysis. The result is that the HIGH binning setting is valuable for quick throughput screening of signal presence and to combine data points to resolve low signal. The LOW binning provides a HIGHER resolution, but takes much longer to process the signal data. The machine’s MEDIUM binning option is a good compromise that is recommended for 90% of all applications.
Field of View
The IVIS Spectrum has 5 possible settings for FOV, denoted by the letters A though E. The camera in the instrument is static, therefore the FOV is changed by having a movable stage. The letters denote the height of the stage, and therefore the proximity (or not) to the camera.
“A” is the closest distance to the camera, and is approximately 2.5cm from the stage at this height. This is ONLY SUITABLE for tissue culture plates and similar, NOT for animal imaging.
As the stage drops to B, C and so on, the FOV widens. This is because the FOV expands like a cone from the lens of the camera, getting proportionally bigger with increasing distance between the stage and camera.
In order to acquire the most data with the best efficiency, it is important to determine the subject height (or maximum subject height if several subjects used). The instrument is set up to reference this value as the height at which to begin scanning. This should be measured and input into the software controls for EVERY data run.