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ImageXpress Confocal HT.ai High-Content Imaging System User Guide
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Cell-Based Assays
The most important consideration when you select cells for a high-content assay is whether
they are compatible with the biology being studied. The assay should give a robust response
with clear distinction between positive and negative phenotypes. It is important to select a
source of cells where it is possible to obtain consistent results from batch to batch, whether
they are primary cells or cell lines, and whether they are transfected or not. Compared to
widefield imaging, confocal imaging provides higher-quality images of thick samples, 3D cell
masses, or tissues. Uniformly staining thick samples for confocal imaging can be difficult.
Organism-Based Assays
You can use the ImageXpress Confocal HT.ai system to image whole organisms, such as
nematodes or zebrafish. Treat the organisms with anesthetic, use a gel preparation, or both to
immobilize the organisms for best results.
Homogeneous Assays
Homogeneous assays are ones where the unbound or unreacted stains are not washed off
before acquiring images. Confocal imaging reduces background fluorescence from out of
focus sample volume which improves signal-to-background ratios and allows for
homogeneous assays that would not work when widefield imaging.
Plates or Slides
Multi-well plates offer several advantages for high-content screening:
The well layout is consistent from one plate to another.
Plates are easier to handle during sample preparation and imaging.
Plates facilitate scaling up to a larger screen.
Some assays, such as imaging of tissue sections, may require the use of slides. A slide holder
is provided in the accessory kit, and there are software tools available to streamline the slide-
imaging workflow.