Mesenchymal stem cells (MSCs) are a multi-potential cell type that can be induced to differentiate to a variety of tissue-specific cell phenotypes, including cartilage (chondrogenesis) and bone (osteogenesis). Given this multi-potentiality, MSCs are a promising cell source for exploring developmental paradigms and for tissue engineering (TE) applications. For cartilage formation assays, MSCs are collected in high-density pellets and treated with specific biofactors, including TGF-β superfamily members and dexamethasone in a chemically defined medium (CM) [1]. During chondrogenesis, extracellular matrix (ECM) rich in glycosaminoglycan (GAG) and type II collagen is synthesized. While MSC chondrogenesis is well-characterized using existing protocols, the effect of alternative biofactors, their doses and combinations requires laborious combinatorial studies [2]. High-throughput screening (HTS) overcomes this limitation through the simultaneous layout and query of a large number of conditions within a single plate. HTS depends on the use of precise robotic liquid handling systems and on the development of sensitive, validated, and readily quantifiable assays. In a recent study, we optimized cell culture and assay procedures for HTS by minimizing cell number, handling and culture duration [3]. We successfully reduced the time scale from 21 to 7 days and the number of cells required from 225K to 30K cells per pellet. Further, we developed a novel in-well digestion protocol to enable high-throughput analysis and minimize handling. In this study, we have further streamlined these assays for HTS by providing a rapid and robotic approach for layout, culture, and analysis of ECM deposition using ‘micro’ MSC pellets (10K cells per pellet) in a 384-well format. Furthermore, we have carried out an initial screen of the NINDS small molecule library and demonstrated the feasibility of this technology for use in HTS of chondrogenesis.
Magnetic Nickel iron Electroformed Trap (MagNET): a master/replica fabrication strategy for ultra-high throughput (>100 mL h−1) immunomagnetic sorting