High Yield Patterning of Single Cells from Extremely Small Populations

Abstract

Many biological assays require the ability to isolate and process single cells. Some research fields, such as the characterization of rare cells, the in vitro processing of stem cells, and the study of early stage cell differentiation, call for the additional and typically unmet ability to work with extremely low-count cell populations. In all these cases, efficient single-cell handling must be matched with the ability to work on a limited number of cells with a low cell loss rate. In this paper, we present a platform combining flow-through processing with deterministic (nonstatistical) patterning of cells coming from extremely small cell populations. We describe here modules using dielectrophoresis to control the position of cells flowing in microchannels and to pattern them in open microwells where cells were further analyzed. K562 cells continuously flowing at a speed of up to 100 μm/s were tridimensionally focused, aligned, and patterned inside microwells. A high-patterning yield and low cell loss rate were demonstrated experimentally: 15uL drops, containing an average of 15 cells, were transferred to the microchannel with an 83% yield, and cells were then patterned into microwells with a 100% yield. The deterministic patterning of cells was demonstrated both by isolating single cells in microwells and by creating clusters composed of a predetermined number of cells. Cell proliferation was assessed by easily recovering cells from open microwells, and a growth rate comparable to the control was obtained.

Negative Dielectrophoretic Capture and Repulsion of Single Cells at a Bipolar Electrode: The Impact of Faradaic Ion Enrichment and Depletion

“…10 Third, DEP has been demonstrated to be useful for single cell manipulation. 12–14 This has been accomplished by constraining the trapping point either by adding physical barriers or by defining an electric field cage similar in size to a single cell. In either case, DEP conditions are chosen that prevent cell–cell attraction, thus discouraging multicell capture.…”

Bioengineering approaches to guide stem cell-based organogenesis

“…In the past, micromanipulation (Jin et al, 2009) and laser microdissection (Revzin et al, 2005) have been used to circumvent this problem, but these approaches are not conducive to automated, parallelized and high-throughput analyses. The recently described inverted microwells, wherein cells are cultured at the air-liquid interface and released on-demand pneumatically (Bocchi et al, 2012;Faenza et al, 2013) are promising alternatives.…”

Analysis of metabolites in single cells—what is the best micro‐platform?

“…Hence, for such systems, high‐density arrays are usually preferred. In the deterministic approach , the cells are guided to their microwells by the use of fluidic flows into the open microwells (i.e. traps).…”

“…Papers of particular interest, published within the period of this review, are mentioned inside the parenthesis. (A) Microfluidic flow‐cytometry platforms ; (B) Microfluidic platforms where cells can be momentarily captured ; (C) Microarray platforms were cells are either fixed or trap for long periods of time .…”