Three-dimensional cell culture microarray for high-throughput studies of stem cell fate.
We have developed a novel three-dimensional (3D) cellular microarray platform to enable the rapid and efficient tracking of stem cell fate and quantification of specific stem cell markers. This platform consists of a miniaturized 3D cell culture array on a functionalized glass slide for spatially addressable high-throughput screening. A microarray spotter was used to deposit cells onto a modified glass surface to yield an array consisting of cells encapsulated in alginate gel spots with volumes as low as 60 nL. A method based on an immunofluorescence technique scaled down to function on a cellular microarray was also used to quantify specific cell marker protein levels in situ. Our results revealed that this platform is suitable for studying the expansion of mouse embryonic stem (ES) cells as they retain their pluripotent and undifferentiated state. We also examined neural commitment of mouse ES cells on the microarray and observed the generation of neuroectodermal precursor cells characterized by expression of the neural marker Sox-1, whose levels were also measured in situ using a GFP reporter system. In addition, the high-throughput capacity of the platform was tested using a dual-slide system that allowed rapid screening of the effects of tretinoin and fibroblast growth factor-4 (FGF-4) on the pluripotency of mouse ES cells. This high-throughput platform is a powerful new tool for investigating cellular mechanisms involved in stem cell expansion and differentiation and provides the basis for rapid identification of signals and conditions that can be used to direct cellular responses.
Fernandes
Tiago G
TG
Department of Chemical and Biological Engineering, Center for Biotechnology and Interdisciplinary Studies, Rensselaer Polytechnic Institute, Troy, New York, USA.
Kwon
Seok-Joon
SJ
Bale
Shyam Sundhar
SS
Lee
Moo-Yeal
MY
Diogo
Maria Margarida
MM
Clark
Douglas S
DS
Cabral
Joaquim M S
JM
Dordick
Jonathan S
JS
D000464:Alginates; D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods; D002454:Cell Differentiation; D049109:Cell Proliferation; D003470:Culture Media / Q000737:chemistry; D051519:Fibroblast Growth Factor 4 / Q000502:physiology; D005782:Gels; D005898:Glass; D051379:Mice; D046228:Microarray Analysis / Q000379:methods*; D055748:SOXB1 Transcription Factors / Q000096:biosynthesis; D013234:Stem Cells
D000464:Alginates; D003470:Culture Media; C496446:Fgf4 protein, mouse; D051519:Fibroblast Growth Factor 4; D005782:Gels; D055748:SOXB1 Transcription Factors; C098693:Sox1 protein, mouse