title: Efficient division and sampling of cell colonies using microcup arrays.
abstract: A microengineered array to sample clonal colonies is described. The cells were cultured on an array of individually releasable elements until the colonies expanded to cover multiple elements. Single elements were released using a laser-based system and collected to sample cells from individual colonies. A greater than an 85% rate in splitting and collecting colonies was achieved using a 3-dimensional cup-like design or "microcup". Surface modification using patterned titanium deposition of the glass substrate improved the stability of microcup adhesion to the glass while enabling minimization of the laser energy for splitting the colonies. Smaller microcup dimensions and slotting the microcup walls reduced the time needed for colonies to expand into multiple microcups. The stem cell colony retained on the array and the collected fraction within released microcups remained undifferentiated and viable. The colony samples were characterized by both reporter gene expression and a destructive assay (PCR) to identify target colonies. The platform is envisioned as a means to rapidly establish cell lines using a destructive assay to identify desired clones.
authors.affiliation: Department of Chemistry, University of North Carolina, Chapel Hill, NC 27599, USA.
authors.forename: Jeng-Hao
authors.initials: JH
authors.lastname: Pai
authors.forename: Kimberly
authors.initials: K
authors.lastname: Kluckman
authors.forename: Dale O
authors.initials: DO
authors.lastname: Cowley
authors.forename: Donna M
authors.initials: DM
authors.lastname: Bortner
authors.forename: Christopher E
authors.initials: CE
authors.lastname: Sims
authors.forename: Nancy L
authors.initials: NL
authors.lastname: Allbritton
mesh_terms: D000818:Animals; D018929:Cell Culture Techniques; D002469:Cell Separation / Q000379:methods*; D053595:Embryonic Stem Cells / Q000166:cytology; D007834:Lasers; D051379:Mice; D055616:Microtechnology; D059010:Single-Cell Analysis; D013997:Time Factors; D046888:Tissue Array Analysis / Q000379:methods*