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Cell Assembly in Flow

Fast Cell Assembly in controlled microfluidic flow

The only way to generate scaffold free spheroids and tissues in minutes.

Microfluidic scheme and images describing the step-wise engineering of cell surfaces in flow followed by cell assembly. (Left) Native cells are flowed through the microfluidic channels and rapidly tailored with bio-orthogonal functional groups via rapid liposome fusion in flow. The tailored cells are then mixed where they assemble into spheroids via an inter cell surface click chemistry reaction. (A) Two cell types engineered with complementary interfacial bio-orthogonal groups. (B) Upon cell to cell contact a rapid multivalent Velcro like click reaction occurs resulting in stable co-culture assembly. (C, D) Changing the engineered cell density ratio injected into the microfluidic device results in spheroids with different morphologies. A 8:1 ratio (GFP:RFP) of cells results in a single GFP NIH 3T3 cell surrounded by RFP Neonatal Dermal Fibroblast cells (C). Reversing the ratio to 1:8 (GFP:RFP) results in a single RFP Neonatal Fibroblast cell surrounded by GFP NIH 3T3 cells (D). (E) Fluorescent image of large 3 cell type spheroid, generated by mixing 1:1:1 ratio of GFP:RFP:Blue C3H10T1/2 stained with CMAC (7-Amino-4- Chloromethylcoumarin) in flow. The three cell type spheroids and tissues were easily generated by engineering GFP and RFP cells with ketones while CMAC stained cells present oxyamine groups. (F) A large spheroid of GFP cells obtained by combining two different populations of GFP cells that present ketone and oxyamine groups. (G) A large spheroid of RFP cells obtained by combining two different populations of RFP cells that present ketone and oxyamine groups. (H) A large spheroid of RFP and GFP cells obtained by flowing a 1:1 ratio of engineered RFP and GFP cells in the microfluidic device. By adjusting the flow rate, cell density and ratio of cell density inputs a range of stoichiometric co-culture spheroid assemblies and size of spheroids may be created.

Complex spheroid and tissue formation in microfluidic flow.

Schematic and images of spheroid and tissue formation in controlled microfluidic flow. An example to show 3 cell type assembly. (A) Using a three inlet microfluidic device, a combination of ketone engineered RFP HNDF cells (cell type 1) and hydroxylamine engineered GFP NIH3T3 (cell type 2) were injected, followed by the addition of CMSO (blue) stained ketone engineered C3H10T1/2 cells (cell type 3) resulted in cell-cell assembly of multiple cell type spheroids through rapid click chemistry. The size of spheroid can be modulated by cell type concentration and flow rate. (B) Direct elution and dilution of the spheroid suspension resulted in three color spheroids (20X) under fluorescence microscopy without the use of trapping agents, polymers, scaffolds or fixation to produce stable live spheroids. (C) Three color spheroids under (40X) fluorescence microscopy demonstrates that RFP HDNF cells are primary adhesive cells driving aggregation with a consistency of roughly 2:1:1 (RFP HDNF: GFP NIH3T3: CMSO C3H10T1/2). (D) 40X confocal microscopy images of concentrated deposition of incubated three cell type tissues generated 3D tissues over 24 hours.

Microfluidic Device and Reagent Partnership

OrganoLinX works collaboratively with industry partners to design and create multiplex microfluidic devices for a range of cell assembly in flow applications.

Learn more about the process of creating custom microfluidic devices, or contact us for more information.