US 12,454,674 B2
Method of encapsulating single cells utilizing an alternating current electrospray
Hsueh-Chia Chang, South Bend, IN (US); Zehao Pan, South Bend, IN (US); Vivek Yadav, South Bend, IN (US); Loan Bui, South Bend, IN (US); and Donny Hanjaya-Putra, South Bend, IN (US)
Assigned to UNIVERSITY OF NOTRE DAME DU LAC, South Bend, IN (US)
Appl. No. 18/039,848
Filed by UNIVERSITY OF NOTRE DAME DU LAC, South Bend, IN (US)
PCT Filed Dec. 3, 2021, PCT No. PCT/US2021/061715
§ 371(c)(1), (2) Date Jun. 1, 2023,
PCT Pub. No. WO2022/120114, PCT Pub. Date Jun. 9, 2022.
Claims priority of provisional application 63/121,369, filed on Dec. 4, 2020.
Prior Publication US 2024/0002781 A1, Jan. 4, 2024
Int. Cl. C12N 5/00 (2006.01); C12N 5/077 (2010.01); C12N 5/09 (2010.01); C12N 13/00 (2006.01)
CPC C12N 5/0012 (2013.01) [C12N 5/0652 (2013.01); C12N 5/0693 (2013.01); C12N 13/00 (2013.01); C12N 2533/30 (2013.01); C12N 2533/54 (2013.01); C12N 2533/74 (2013.01); C12N 2537/10 (2013.01)] 20 Claims
OG exemplary drawing
 
1. A method for high-throughput single-cell encapsulation and tip streaming, the method comprising:
(a) combining a plurality of cells with a hydrogel and oil to generate a water-in-oil emulsion;
(b) agitating the emulsion to drive single cells into the center of individual hydrogel beads to generate single-cell encapsulated hydrogel beads;
(c) crosslinking the hydrogel;
(d) extracting the crosslinked single-cell encapsulated hydrogel beads from the emulsion into cell media;
(e) transferring the extracted single-cell encapsulated hydrogel beads into a micropipette;
(f) applying a back pressure and alternating current (AC) electric potential to the micropipette to generate an AC Taylor cone encompassing a droplet containing the single-cell encapsulated hydrogel bead at the micropipette tip; and
(g) ejecting the droplet containing the single-cell encapsulated hydrogel bead from the micropipette tip.