Fig 1.
Microfluidic single-cell processing platform and experimental workflow.
(A) The microfluidic device consists of cell inlet for delivering cells into the chambers, medium inlet for supplying fresh medium into the chambers after trapping single-cells, 8 individually addressable proliferation chambers, (B) a valve-based junction for the separation of the sister cells after division with a feedback channel that allows relocation of the sister cells after division from the separation area to the cell trapping chambers and (C) extraction wells for the collection of the sister cells. The workflow of the device comprises trapping of a single-cell inside a growth chamber, cell growth and division, separation of the sister cells after division and extraction of the individual sister cells for downstream transcriptome analysis.
Fig 2.
Single-cell proliferation experiments.
(A) Single 6C2 cells, (B) Single T2EC cells were trapped and monitored over a period of 24 hours. Time lapse brightfield images for one chamber were acquired in every 5 minutes. The brightfield images show that cell full division occurs in each chamber within 20 minutes. The scale bar is 50 μm.
Fig 3.
Single T2EC cell proliferation and sister cell relocation.
A single T2EC cell was trapped and monitored by time-lapse brightfield microscopy, with images being acquired every 5 minutes. During sister cell relocation, the first sister is kept in the initial chamber, with the second sister being moved in a new chamber, by applying 10 mbar of pressure from the medium inlet which allows precise control of the single-cell movement. The scale bar is 50 μm.
Fig 4.
(A) A single 6C2 sister cell is monitored using fluorescence imaging in the extraction well. Manual extraction of this cell is performed using a small glass capillary. (B) Fluorescence images of the extraction well before and after the extraction of a single sister cell. The scale bars are 100 μm.
Fig 5.
(A) Plot of the ratio of ERCC mapped in each cell. The orange line represents the cut off value; cells positioned higher than the cut off are discarded. (B) Boxplot showing the number of detected genes per cell, sorted with conventional FACS or cultured and isolated using the microfluidic platform. A Wilcoxon rank-test was performed. (C) Boxplot of log(UMIs) number per cell sorted with conventional FACS or cultured and isolated using the microfluidic platform. A Wilcoxon rank-test was performed. (D) UMAP dimensions reduction and projection of the cells. Only complete couples of sister cells were kept for the analysis. Chip-cultured cells are coloured and grouped by lineage and FACS sorted cells are grey.