title: ALISSA: an automated live-cell imaging system for signal transduction analyses.
abstract: Probe photobleaching and a specimen's sensitivity to phototoxicity severely limit the number of possible excitation cycles in time-lapse fluorescent microscopy experiments. Consequently, when a study of cellular processes requires measurements over hours or days, temporal resolution is limited, and spontaneous or rapid events may be missed, thus limiting conclusions about transduction events. We have developed ALISSA, a design framework and reference implementation for an automated live-cell imaging system for signal transduction analysis. It allows an adaptation of image modalities and laser resources tailored to the biological process, and thereby extends temporal resolution from minutes to seconds. The system employs online image analysis to detect cellular events that are then used to exercise microscope control. It consists of a reusable image analysis software for cell segmentation, tracking, and time series extraction, and a measurement-specific process control software that can be easily adapted to various biological settings. We have applied the ALISSA framework to the analysis of apoptosis as a demonstration case for slow onset and rapid execution signaling. The demonstration provides a clear proof-of-concept for ALISSA, and offers guidelines for its application in a broad spectrum of signal transduction studies.
authors.affiliation: Department of Physiology & Medical Physics, Royal College of Surgeons in Ireland, Dublin, Ireland.
authors.forename: Jakub
authors.initials: J
authors.lastname: Wenus
authors.forename: Heiko
authors.initials: H
authors.lastname: Düssmann
authors.forename: Perrine
authors.initials: P
authors.lastname: Paul
authors.forename: Dimitrios
authors.initials: D
authors.lastname: Kalamatianos
authors.forename: Markus
authors.initials: M
authors.lastname: Rehm
authors.forename: Peter
authors.initials: P
authors.lastname: Wellstead
authors.forename: Jochen
authors.initials: J
authors.lastname: Prehn
authors.forename: Heinrich
authors.initials: H
authors.lastname: Huber
mesh_terms: D017209:Apoptosis; D001331:Automation; D002460:Cell Line; D002470:Cell Survival; D002477:Cells / Q000166:cytology* / Q000378:metabolism*; D003196:Computer Graphics; D031541:Fluorescence Resonance Energy Transfer; D021621:Imaging, Three-Dimensional / Q000379:methods*; D008928:Mitochondria / Q000378:metabolism; D051336:Mitochondrial Membranes / Q000378:metabolism; D009862:Online Systems; D015398:Signal Transduction; D012984:Software; D013997:Time Factors; D014584:User-Computer Interface