IC200: Running A Plate

This lesson describes how to run a plate.

Create a new project via the 'File --> New Project' command.

Define an experiment name, select the plate type and set a location to save files. A new directory will be created for each plate.

A new project window will load. Select the algorithm, region within each well to scan, channel settings, KIC Details, wells to scan and then start the scan.

Select wells to scan. Every well highlighted in blue will be scanned. This can be redone at any point before selecting 'Start Scanning.'

Select the algorithm protocol to scan. Algorithm protocols can be edited and created via 'File --> Edit Algorithms...'

Editing algorithms is covered in another lesson.

Select the image fields to scan within each well.

Adjust settings for each channel. Light source, intensity, time, gain and binning can be adjusted here. Default values can be adjusted via the algorithm protocol editing tool.

- The light source is a selection from whichever light sources are available to the system.
- The intensity of the light source has a discrete range of zero to 255, with 255 being the brightest setting.
- Time is indicated in milliseconds.
- The range for valid gain values varies from system to system. When is doubt, a value of '1' generally works well.
- Binning can be set here if available on the system. Values of 1,2,4 or 8 are sometimes referred to 1x1, 2x2, 4x4 and 8x8 binning. The two notations are equivalent.

- Optionally, click 'Snap' in the lower left hand corner to acquire updated images with the given settings. This assumes a plate is already loaded onto the system.

The result of a 'Snap' generates images for each color in your scan given the settings shown. It is acquired from the first well - upper left - selected from your well plate.

Tip:

- Select the 'Auto-Contrast' checkbox for more routine image viewing.
- Select between tabs to view the images from each channel.
- Select the 'Histogram' checkbox to view the histogram of the image in the current tab.

A log-linear histogram view is an effective tool to estimate the use of the dynamic range of a given image.

This table describes the standard excitation/emission set. The (colors) in parenthesis correspond to the colors in the 'Channel Settings' dialog. The numbered sequence indicates the standard filter wheel position associated with each excitation. Some fluorophores may require custom matching of excitation light source and emission filter wheel position.

Some system configurations may vary.