US 12,345,701 B2
Methods for assaying biological cells in a microfluidic device
Yelena Bronevetsky, Emeryville, CA (US); Annamaria Mocciaro, Emeryville, CA (US); Guido K. Stadler, Emeryville, CA (US); Peter J. Beemiller, Emeryville, CA (US); Natalie C. Marks, Emeryville, CA (US); Duane Smith, Emeryville, CA (US); Vincent Haw Tien Pai, Emeryville, CA (US); Jason M. McEwen, Emeryville, CA (US); Amanda L. Goodsell, Emeryville, CA (US); John A. Tenney, Emeryville, CA (US); Thomas M. Vetterli, Emeryville, CA (US); and Hansohl E. Kim, Emeryville, CA (US)
Assigned to BRUKER CELLULAR ANALYSIC, INC., Emeryville, CA (US)
Filed by BRUKER CELLULAR ANALYSIS, INC., Emeryville, CA (US)
Filed on Apr. 27, 2021, as Appl. No. 17/241,865.
Application 17/241,865 is a continuation of application No. PCT/US2019/059495, filed on Nov. 2, 2019.
Claims priority of provisional application 62/754,107, filed on Nov. 1, 2018.
Claims priority of provisional application 62/754,147, filed on Nov. 1, 2018.
Claims priority of provisional application 62/881,129, filed on Jul. 31, 2019.
Prior Publication US 2021/0349075 A1, Nov. 11, 2021
Int. Cl. G01N 33/50 (2006.01); B01L 3/00 (2006.01)
CPC G01N 33/505 (2013.01) [B01L 3/502761 (2013.01); G01N 33/5014 (2013.01); B01L 2200/0647 (2013.01)] 23 Claims
 
1. A method of assaying for antigen-specific cytotoxicity of a T lymphocyte (T cell) in a microfluidic device, the method comprising:
disposing the T cell within the microfluidic device; wherein the microfluidic device comprises a flow region for containing a flow of a first fluidic medium and a chamber opening to the flow region;
disposing a target cell in proximity to the T cell, wherein the target cell is configured to capture a secreted biomolecule from the T cell;
detecting the secreted biomolecule captured by the target cell after a period of exposure in proximity to the T cell by:
receiving a first image and one or more second images of a region of interest;
transforming each of the one or more second images to optically align with the first image;
processing pixel data in the first image using a machine learning algorithm to detect target cells present in the region of interest, the detection comprising identifying a boundary of each target cell; and
detecting a signal located within the boundary of each detected target cell in each of the one or more second images; and
determining the viability of the target cell after a period of exposure in proximity to the T cell.