Papers & Publications

Publications Using the CellRaft® Technology

See these scientific publications for demonstrations of the various uses for products based on the CellRaft® Technology.

For identifying potential aneuploidy in normal somatic tissues, techniques such as fluorescence in situ hybridization (FISH) and single-cell low-coverage whole genome sequencing (scL-WGS) have been utilized. The accuracy of these techniques including prevalence for false positive/negative accounts of aneuploidy have been inconsistent. To understand the efficiency of these techniques, cells with naturally occurring or induced aneuploidy were generated and isolated with the CellRaft single cell picking system. After CellRaft isolation, these single cells were amplified and sequenced with scL-WGS. The results showed that scL-WGS frequently underestimated aneuploidy levels while FISH overestimated, and a modified 2-probe approach can be used as additional detection for low levels of aneuploidy.

CellRaft Arrays were used as a culture device for seeding gastrointestinal organoid fragments for studying the effects of microbiota. Using a custom made microinjection system, E. coli was injected into the organoids housed on the CellRaft Array and they were imaged to insure proper injection. The Quad Array allowed for 4 different media chambers to test for different conditions and various antibiotics. The use of this customized hardware modification in conjunction with the CellRaft Array allowed for the development of a semi-automated high-throughput organoid microinjection system that could be reproduced in many laboratories.

Gracz AD, Williamson IA, Roche KC, Johnston MJ, Wang F, Wang Y, Attayek PJ, Balowski J, Liu XF, Laurenza RJ, Gaynor LT, Sims CE, Galanko JA, Li L, Allbritton NL, Magness ST
Nat Cell Biol. 2015 Mar;17(3):340-9. doi: 10.1038/ncb3104. Epub 2015 Feb 9

In order to determine the underlying mechanisms of a broad range of issues related to human health and disease, it is important to first understand how somatic stem cells self-renew and differentiate to produce the functional cells of the resident tissue, as stem cells reside in niches where support cells provide signaling critical for tissue renewal. Magness and colleagues demonstrated the use of the CellRaft® Technology to prove that Paneth cells (PC), a known intestinal stem cell (ISC) niche component, enhance organoid formation in a contact-dependent manner. CellRafts® were used to facilitate retrieval of early enteroids for qPCR to correlate functional properties, such as enteroid morphology, with differences in gene expression. This platform enabled the study of a large number of single ISCs simultaneously, either at the clonal level or in the presence of niche cells, with multi-day, three-dimensional culture in extracellular matrices applied directly to the CytoSort™ Arrays. The authors found that direct cell-to-cell contact between ISCs and PCs is required for enhanced ISC growth.

Currently, people with HIV (PWH) are dependent on uninterrupted regimens of antiretroviral therapy (ART) as reservoirs of latent infected cells remain despite treatment and can cause a rebound if ART is discontinued. Measuring the size of the HIV latent reservoir is useful in determining the effectiveness of cures that silence or eliminate these reservoirs, but current methods are imprecise, challenging, and time-consuming.

In this article, the authors describe a novel technique for identifying the size of the latent infected cell reservoir in PWH using an HIV-detecting reporter cell line and Cell Microsystem’s CellRaft technology, which they refer to as the Microwell Outgrowth Assay (MOA). In this assay, CellRaft Arrays allow for co-culturing of their fluorescent reporter cell line and CD4 T cells from PWH, while the CellRaft AIR System and software can automatically image and identify rare events such as reporter cells that fluoresce when infected. These infected cells can also be automatically isolated for downstream analysis and viral sequencing. Thus, not only is MOA a quick and scalable method for measuring viral outgrowth, but also can provide valuable information for HIV cures.

Matthew Simon, Jiping Yang, Jonathan Gigas, Eric J Earley, Eric Hillpot, Lei Zhang, Maria Zagorulya, Greg Tombline, Michael Gilbert, Samantha L Yuen, Alexis Pope, Michael Van Meter, Stephan Emmrich, Denis Firsanov, Advait Athreya, Seyed Ali Biashad, Jeehae Han, Seungjin Ryu, Archana Tare, Yizhou Zhu, Adam Hudgins, Gil Atzmon, Nir Barzilai, Aaron Wolfe, Kelsey Moody, Benjamin A Garcia, David D Thomas, Paul D Robbins, Jan Vijg, Andrei Seluanov, Yousin Suh, and Vera Gorbunova
The EMBO Journal – October 2022

Sirtuin 6 (SIRT6) is a deacylase and mono-ADP-ribosyltransferase enzyme implicated in multiple aging-related pathways including DNA repair, telomere maintenance, and metabolism regulation. A strong correlation between SIRT6 activity and longevity has been observed across species, and genetic polymorphisms in the SIRT6 gene correlate with human longevity in SNP analyses. To demonstrate that the centSIRT6 allele improves DNA repair efficiency, the authors used the CellRaft AIR system to isolate monoclonal mesenchymal stem cells (MSCs) with a centSIRT6 knock-in. MSCs with the centSIRT6 allele demonstrated a higher efficiency of DNA repair and increased survival upon treatment with methyl methanesul-fonate, a potent DNA-damaging agent, supporting the hypothesis that the centSIRT6 allele contributes to human longevity by improving genome maintenance.

Most of the human genome is transcribed but only a small percentage of the transcripts are translated. The most abundant non-translated transcripts are long non-coding RNAs (IncRNAs). This review aims to collect updated information about IncRNA classification and new findings on their function derived from single-cell analysis. The method described here involves using CellRaft Arrays to seed individual cells of interest and then isolate them into PCR tubes or plates for further analysis and sequencing. This review describes the process as “minimal disturbance on cells during the capture and is optimized for adherent cells.”

Raft Notes