Microfluidic Enrichment of Mouse Epidermal Stem Cells and Validation of Stem Cell Proliferation In Vitro

Abstract

Bulge stem cells reside in the lowest permanent portion of hair follicles and are responsible for the renewal of these follicles along with the repair of the epidermis during wound healing. These cells are identified by surface expression of CD34 and the a6-integrin. When CD34 and a6 double-positive cells are isolated and implanted into murine skin, they give rise to epidermis and hair follicle structures. The current gold standard for isolation of these stem cells is fluorescence-activated cell sorting (FACS) based on cell surface markers. Here, we describe an alternative method for CD34 bulge stem cell isolation: a microfluidic platform that captures stem cells based on cell surface markers. This method is relatively fast, requiring 30 min of time from direct introduction of murine skin tissue digestate into a two-stage microfluidic device to one-pass elution of CD34+ enriched cells with a purity of 55.8% -5.1%. The recovered cells remain viable and formed colonies with characteristic morphologies. When grown in culture, enriched cells contain a larger a6 + population than un-enriched cells.

Parametric control of collision rates and capture rates in geometrically enhanced differential immunocapture (GEDI) microfluidic devices for rare cell capture

“…Recent work has successfully isolated CD34-positive endothelial stem (Zhu et al, 2013) and progenitor cells (Hatch et al, 2011), and CTCs from cancer patients (Hsieh et al, 2006; Nagrath et al, 2007; Talasaz et al, 2009; Stott et al, 2010; Gleghorn et al, 2010) with microfluidic devices. These devices can be divided into two major categories: pressure-driven devices and those that also use electrokinetic techniques (primarily dielectrophoresis).…”

A transfer function approach for predicting rare cell capture microdevice performance

“…Unless otherwise noted, the results described in this work are for an array with row spacing = 200 μm, column spacing = 200 μm, obstacle radius R = 50 μm, an array length of N = 100 unit structures, and U inlet = 100 μms; this geometry and flow rate corresponds to those used in previously reported experiments (Gleghorn et al 2010;Zhu et al 2013). Results are presented in dimensional units to facilitate ready comparison to biological length scales.…”

“…Fetal cells circulating in maternal blood present a non-invasive alternative to amniocentesis (Mohamed et al 2007), reducing the risk to the fetus. Stem (Zhu et al 2013) and progenitor cells (Hatch et al 2011) have been used for culture and later experimentation, including transplantation . Circulating epithelial cells (CECs) of pancreatic origin have been found in the blood of pancreatic cyst patients, and may enable the early detection of pancreatic cancer .…”

Fundamentals and application of magnetic particles in cell isolation and enrichment: a review

“…Briefly, microfluidic channels can be functionalized with affinity biomolecules and, similar to the initial work by Wigzell et al (Wigzell and Andersson, 1969), cells can be selectively captured from a flow channel (Didar and Tabrizian, 2010). Numerous examples exist in the literature that have illustrated the effectiveness of microfluidic cell affinity chromatography for the isolation of circulating tumor cells (Gleghorn et al, 2010, Stott et al, 2010a, Nagrath et al, 2007, Adams et al, 2008a, Du et al, 2006), endothelial progenitor cells (Hansmann et al, 2011, Plouffe et al, 2009b, Hatch et al, 2011, Hatch et al, 2012), endothelial and smooth muscle cells (Plouffe et al, 2009b, Green and Murthy, 2009, Plouffe et al, 2007, Plouffe et al, 2008), skin stem cells (Zhu et al, 2013), white blood cells (Murthy et al, 2004, Sin et al, 2005, Xu et al, 2009). Although microfluidic capture channels have illustrated excellent recoveries (> 90%) and purities (> 95%) (Didar and Tabrizian, 2010) of very rare cells versus many alternative approaches the difficulty in gently removing trapped cells from the surface of the affinity substrate has limited the use in many biological fields (Murthy and Radisic, 2008).…”