Laser-scanning imaging (LSI) as employed by the acumen® eX3 (TTP Labtech) complements CCD-based HCS instruments, providing rapid “on the fly” analysis of multiplexed assays.
In this system, in place of a microscope, a scanning laser beam is passed through an F theta lens, enabling rapid analysis over a large field of view and depth of field. Emitted fluorescence from labelled cells is detected via a series of photomultiplier tubes (PMT), providing a 2D image similar to that generated by a CCD camera.
Serial PMTs enable simultaneous four-color fluorescence detection for multiplexing studies enabling high-content information to be calculated for every object. High-content data can be exported to proprietary or open-source software, allowing the system to address a range of applications routinely requiring a 20x objective using CCD-based technologies.
Although lacking the higher optical resolutions of the more costly CCD-based imagers, acumen’s widefield objective provides whole well imaging capabilities enabling the capture and positional analysis of every cell in a single scan. This feature eliminates the need for multiple image capture to achieve suitable cell numbers for data normalization and overcomes issues of uneven cell distribution across a well.
In addition, it is possible to filter cell debris or fluorophore crystals based on size, to provide statistically robust data from every well. Whole-well imaging and simultaneous collection of multiparametric data eliminates the need to process multiple images or colors in each well resulting in small files sizes (50KB CSV), which can be easily transported to screening databases. In screening mode, acumen allows validated assays to be routinely and rapidly carried out without the need for detailed image processing and large data file storage.
acumen’s large depth of field also enables high-content image analysis of cells, spheroids and cell colonies, or objects growing on within or a gel for the study of a range of applications such as cell-cycle analysis, cell migration, and live-cell studies, as well as automated analysis of tissues and small organisms. This attribute overcomes the additional imaging times required following refocusing and reimaging using CCD-based imagers or the restricted depth of field imposed by microscopes.
Laser-scanning imaging technology enables rapid plate scanning and analysis independent of plate size (from 96 to 1,536 wells). With an acquisition rate up to 40 images per second, assay miniaturization and minimal cell culture usage, the system saves valuable sample and reagent costs without impacting assay throughput.
With “on the fly” data acquisition and analysis, acumen is an ideal instrument for HCS of compound libraries where rapid data output is required.