Automated Profiling of Individual Cell-Cell Interactions from High-throughput Time-lapse Imaging Microscopy in Nanowell Grids (TIMING)
This page describes the FARSIGHT approach of automated profiling of individual cell-cell interactions applied to time-lapse imaging microscopy in nanowell grids.
Contents |
Background and Motivation
There is a need for effective automated methods for profiling dynamic cell-cell interactions with single-cell resolution from high-throughput time-lapse imaging data, especially, the interactions between immune effector cells and tumor cells in adoptive immunotherapy.
Fluorescently labeled human T cells, Natural Killer cells (NK), and various target cells (NALM6, K562, EL4) were co-incubated on PDMS arrays of sub-nanoliter wells (nanowells), and imaged using multi-channel time-lapse microscopy. The proposed cell segmentation and tracking algorithms account for cell variability and exploit the nanowell confinement property to increase the yield of correctly analyzed nanowells from 45% (existing algorithms) to 98% for wells containing one effector and a single target, enabling automated quantification of cell locations, morphologies, movements, interactions, and deaths without the need for manual proofreading. Automated analysis of recordings from 12 different experiments demonstrated automated nanowell delineation accuracy >99%, automated cell segmentation accuracy >95%, and automated cell tracking accuracy of 90%, with default parameters, despite variations in illumination, staining, imaging noise, cell morphology, and cell clustering. An example analysis revealed that NK cells efficiently discriminate between live and dead targets by altering the duration of conjugation. The data also demonstrated that cytotoxic cells display higher motility than non-killers, both before and during contact.
Algorithm Pipeline
Supplementary Materials
Software and Test Data
https://github.com/lvyb/F_nanowell/tree/master/software https://github.com/lvyb/F_nanowell/tree/master/test%20data
Tracking Algorithm Comparison
https://github.com/lvyb/F_nanowell/tree/master/tracking%20comparison
