Enrichment and expansion of cells using antibody‐coated micropallet arrays

Abstract

Positive selection, sorting, and collection of single cells from within a heterogeneous population are required for many biological studies. We recently demonstrated a miniaturized cell array for this purpose; however, on-chip pre-enrichment and isolation of specific target cells would provide significant value for cell isolation. In the current work, mixed cell samples of fewer than 30,000 cells were used for panning by means of on-array antibody-capture to pre-enrich the target population. The cell surface receptors Fc e R 1 , c-Kit, and ErbB2 were used for positive selection of RBL, RBL, and SK-BR-3 cells, respectively, from the mixed population. The capture efficiency, selectivity, and enrichment for the target cells were calculated and compared with fibronectin-coated controls. As expected, the capture efficiency depended on the frequency of the target cell in the mixed population over the range of 0.3-33%. For a frequency of 5% target cells, the capture efficiency was 39%-53% for the three conditions, while the selectivity varied between 78% and 98% with 16-20-fold enrichment. Furthermore, single-cell cloning studies demonstrated a high cloning efficiency of target cells selectively isolated from the array. Antibody-based pre-enrichment in combination with micropallet-based cell selection will be a valuable tool for isolation and expansion of rare cells from small heterogeneous populations. '

Polystyrene-coated micropallets for culture and separation of primary muscle cells

“…The 1002F photoresist from which the micropallet arrays were fabricated has been shown to be suitable for culturing tumor cell lines [15,21]. In some instances, the 1002F required an additional coating such as collagen or fibronectin for cell attachment and growth.…”

“…Contact printing of the micropallets was evaluated for its capacity to generate a suitable surface for the culture of PCSCs and to lay the groundwork for developing procedures applicable to other primary cell types [19–21]. The long-term goal of this work is to utilize the micropallet arrays to sort PCSCs using a variety of parameters and shorter timescales not available through traditional cloning techniques or flow cytometry.…”

Large area magnetic micropallet arrays for cell colony sorting

“…Previous work has shown that extracellular matrix proteins and specific antibodies have a significant effect on the retention of cells to microstructures fabricated from 1002F photoresist. 21,22 LabTek 8-well polystyrene cell culture chamber slides (Nunc, Naperville, IL) were mounted to the surface of the microarrays with polyIJdimethylsiloxane) and placed in an oven at 65 °C for 15 minutes, solidifying the adhesive and creating a liquid seal. The arrays were sterilized in 70% ethanol and dehydrated in a sterile environment.…”

Highly efficient cellular cloning using Ferro-core Micropallet Arrays

“…Shadpour et al . reported the ability to identify colonies derived from murine ES cells by gross morphology using mechanical roughening of the surface, that was required for the incorporation of a gelatin substrate 16,17 and the coating of the micropallet array with a monoclonal antibody recognizing a cell surface molecule 18 , a functionalization of the micropallet array that has been used to capture rare cells from heterogeneous mixtures 10 . We described the ability to effectively coat the micropallet array with a variety of extracellular matrix components that could be tailored to the desired adherent cell type 19 , including methodologies to limit bridging of the extracellular matrix coating between micropallets.…”