Platforms for Single-Cell Collection and Analysis

Abstract

Single-cell analysis has become an established method to study cell heterogeneity and for rare cell characterization. Despite the high cost and technical constraints, applications are increasing every year in all fields of biology. Following the trend, there is a tremendous development of tools for single-cell analysis, especially in the RNA sequencing field. Every improvement increases sensitivity and throughput. Collecting a large amount of data also stimulates the development of new approaches for bioinformatic analysis and interpretation. However, the essential requirement for any analysis is the collection of single cells of high quality. The single-cell isolation must be fast, effective, and gentle to maintain the native expression profiles. Classical methods for single-cell isolation are micromanipulation, microdissection, and fluorescence-activated cell sorting (FACS). In the last decade several new and highly efficient approaches have been developed, which not just supplement but may fully replace the traditional ones. These new techniques are based on microfluidic chips, droplets, micro-well plates, and automatic collection of cells using capillaries, magnets, an electric field, or a punching probe. In this review we summarize the current methods and developments in this field. We discuss the advantages of the different commercially available platforms and their applicability, and also provide remarks on future developments.

Guidelines for the use of flow cytometry and cell sorting in immunological studies (second edition)

“…Especially the invention of commercially available systems from 10× Genomics, Dolomite Bio, and 1Cell Bio, made these technologies broadly available. Nanowell‐based systems process thousands of cells in a single step by depositing barcoded beads and cells into nanowells on a printed chip . These systems are commercially available, e. g., from BD Biosciences (Rhapsody), Fluidigm (C1), or Clontech (ICell8).…”

A Single Cell but Many Different Transcripts: A Journey into the World of Long Non-Coding RNAs

“…Depending on the biological question to be addressed and on the type of starting material, a pool of cells can be enriched based on its particular morphophysiological features (e.g., size, shape, electrical charge, expression of molecular markers). Different technologies can be used for enrichment, such as microfiltration, density gradient centrifugation, immunoaffinity, and magnetic-activated cell sorting [112]. However, the major issue of pre-analytical processing is the disturbance of native expression profiles of cells which can lead to misinterpretation of data.…”

“…Other instruments (e.g., Chromium System-10× Genomics, Pleasanton, CA, USA; InDrop System-1CellBio, Watertown, MA, USA; C1 System-Fluidigm, San Francisco, CA, USA; ddSEQ Single-Cell Isolator-Bio-Rad-Illumina, Hercules, CA, USA) can perform the reactions inside the droplet, increasing the efficiency of the method. These platforms enable the analysis of up to 10,000 single cells (except the C1 System that process up to 800 cells) in a parallel manner, avoiding cross-contamination and increasing reproducibility [112]. However, the RNA capturing efficiency of these techniques can reach 65% maximum [112] and they require high input and clean samples because the microfluidic channels can clog if there are cell clumps or debris.…”

“…These platforms enable the analysis of up to 10,000 single cells (except the C1 System that process up to 800 cells) in a parallel manner, avoiding cross-contamination and increasing reproducibility [112]. However, the RNA capturing efficiency of these techniques can reach 65% maximum [112] and they require high input and clean samples because the microfluidic channels can clog if there are cell clumps or debris. The current trend is to use instruments that provide end-to-end solutions, such as the C1 System and the Chromium System, that process from the single-cell isolation to data analysis in a single platform.…”

VyCAP's puncher technology for single cell identification, isolation, and analysis

“…Over the last couple of decades, multiple technologies have been developed for the enrichment and isolation of rare cells, including circulating tumor cell (CTC). Compatibility between these technologies, is however mostly not present . Enriched sample volumes are too large or the number of unwanted cells in the enriched samples is too high to identify and isolate the cells of interest.…”