Single-cell isolation allows genomic and transcriptomic analysis of one individual cell. It is also required to build a monoclonal cell line from rare cell isolation, that could be for example CRISPR-CAS9 gene edited cells. There are 3 popular methods: serial dilution, micro-manipulation and flow cytometry. None of these are easy or simple, and they often require expertise and experience. Fortunately, a new solution for everybody has come! It’s designed to isolate a single cell in a few seconds, and it’s (quite appropriately) called the Smart Aliquotor.
Very often primary cell cultures are jeopardized by the outgrowth of contaminating fibroblasts. The population of the target primary cells is directly affected since fibroblasts usually grow at much faster rates than other cell types.
Functionality and viability of primary cells can be impaired by incorrect thawing procedures, storage or culture conditions. It’s generally admitted that applying the same protocols as for cell lines leads will lead to bad cell quality. Well, based on our experience with primary cells, here are a few tips you can follow to ensure you get the best performance.
Vector-free CRISPR-CAS9 gene editing to accelerate therapeutic applications
A few years ago, Ayal Hendel et al (doi:10.1038/nbt.3290) published results revealing that chemical alterations to sgRNA enhance gene editing in primary cells. To demonstrate this, Matthew H Porteus’s team chose the targeted genes CCR5, HBB and IL2RG respectively involved in anti-HIV clinical trials, cell anaemia and thalassemia, and severe combined immunodeficiency. More recently, the same team tested several modified CAS9 mRNA. You can find the practical results on this poster introduced at the ASGC.
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