	US 12,241,888 B2
	Platform for assessment of the transcytosis of extracellular vesicles across endothelial barriers and uses thereof
Marsha A. Moses, Brookline, MA (US); and Golnaz Morad, Boston, MA (US)
Assigned to Children's Medical Center Corporation, Boston, MA (US)
Appl. No. 16/979,710
Filed by Children's Medical Center Corporation, Boston, MA (US)
PCT Filed Mar. 14, 2019, PCT No. PCT/US2019/022194 § 371(c)(1), (2) Date Sep. 10, 2020, PCT Pub. No. WO2019/178317, PCT Pub. Date Sep. 19, 2019.
Claims priority of provisional application 62/643,145, filed on Mar. 14, 2018.
Prior Publication US 2021/0041421 A1, Feb. 11, 2021
Int. Cl. G01N 33/50 (2006.01); A61K 31/4709 (2006.01); A61K 31/517 (2006.01); G01N 33/574 (2006.01)

CPC G01N 33/5064 (2013.01) [A61K 31/4709 (2013.01); A61K 31/517 (2013.01); G01N 33/57415 (2013.01); G01N 33/57419 (2013.01); G01N 33/57423 (2013.01); G01N 33/5743 (2013.01); G01N 33/57438 (2013.01)]

21 Claims

1. A method of quantifying transcytosis of an extracellular vesicle (EV) across the blood brain barrier (BBB), the method comprising:

(i) contacting the EV with a layer of brain endothelial cells cultured on a permeable cell culture insert that divides a cell culture well into an upper chamber and a lower chamber, wherein the contacting of the EV with the layer of brain endothelial cells occurs in one of the upper chamber and the lower chamber, and wherein the brain endothelial cells are treated with an agent that increases intracellular cyclic AMP (CAMP) level;

(ii) detecting an EV signal in the other one of the upper chamber and the lower chamber; and

(iii) quantifying the EV signal detected in (ii) from EVs that crossed the layer of brain endothelial cells via transcytosis, wherein the EV signal is higher than a control signal.