EB sorting (Peter Lillehoj)

EB sorting

(Peter Lillehoj)

With their developmental potential to differentiate into all three of the germ layers (endoderm, mesoderm and
ectoderm), embryonic stem (ES) cells provide a unique opportunity to study lineage commitment and can potentially
serve as a source of specialized cells for regenerative medicine. Among the various published ES cell differentiation
protocols, the formation of embryoid bodies (EBs), spherical aggregates of spontaneously differentiating ES cells,
is commonly utilized as a critical intermediate step. EBs appear to recapitulate embryonic development, facilitating
induction of differentiation and commitment into specific cell types and obtaining EBs of homogeneous size appears
to be a key factor for successful ES cell research.

Current pipetting methods for separating EBs are time consuming, inefficient and result in poor size uniformity.
Additionally, the separation of EBs through external force fields, such as dielectrophoretic (DEP), acoustic or
magnetic, may raise potential issues in damaging the fragile cellular entities, thereby affecting subsequent cell
differentiation. To overcome these obstacles, we have developed a microfluidic device for sorting embryoid bodies
(EBs) with large dynamic size ranges up to 300 μm. The proposed separation scheme utilizes appropriately spaced
pillars within a microchannel to alter the fluid flow pathway, thus allowing particles of defined sizes to be diverted
towards specific flow paths. We demonstrate for the first time on-chip separation of mouse EBs, which were
separated into three size groups with separation efficiencies approaching 80%. The ability to extract specific size
ranges of EBs will greatly facilitate their subsequent differentiation studies.