True monolayer cell culture in a confined 3D microenvironment enables lineage informatics.
BACKGROUND
There is a need for methods to (1) track cells continuously to generate lineage trees; (2) culture cells in in vivo-like microenvironments; and (3) measure many biological parameters simultaneously and noninvasively. Herein, we present a novel imaging culture chamber that facilitates "lineage informatics," a lineage-centric approach to cytomics.


METHODS
We cultured cells in a confined monolayer using a novel "gap chamber" that produces images with confocal-like qualities using standard DIC microscopy. Lineage and other cytometric data were semiautomatically extracted from image sets of neural stem and progenitor cells and analyzed using lineage informatics.


RESULTS
Cells imaged in the chamber every 3 min could be tracked for at least 6 generations allowing for the construction of extensive lineage trees with multiparameter data sets at hundreds of time points for each cell. The lineage informatics approach reveals relationships between lineage, phenotype, and microenvironment. Mass transfer characteristics and 3D geometry make the chamber more in vivo-like than traditional culture systems.


CONCLUSIONS
The gap chamber allows cells to be cultured, imaged, and tracked in true monolayers permitting detailed informatics analysis of cell lineage, phenotype, and fate determinants. The chamber is biomimetic and straightforward to build and use, and should find many applications in long-term cell imaging.
Ramunas
John
J
Department of Chemical Engineering, University of Waterloo, Waterloo, Ontario N2L 3G1, Canada.
Illman
Meredith
M
Kam
Angela
A
Farn
Kristen
K
Kelly
Liam
L
Morshead
Cindi M
CM
Jervis
Eric J
EJ
D000818:Animals; D018929:Cell Culture Techniques / Q000295:instrumentation / Q000379:methods*; D002454:Cell Differentiation; D019070:Cell Lineage; D002478:Cells, Cultured; D016895:Culture Media, Serum-Free; D048088:Informatics / Q000295:instrumentation / Q000379:methods*; D008297:Male; D051379:Mice; D013234:Stem Cells / Q000502:physiology
D016895:Culture Media, Serum-Free