- Sony Biotechnology
- Sony Biotechnology
Getting the most from your cell sorter Fluorescence-activated cell sorting (FACS) is an increasingly common step in many life science research applications, from sequencing to protein design to immunotherapy. No matter which downstream application is used, it is possible to take steps to optimize the purity, efficiency, and reliability of each sort performed.
Recent advances in genome editing and the application of fluorescent proteins have accelerated Interest in isolating specific populations of brain cells from mixed populations. Researchers are also using cell sorting to isolate single cells for expansion and analysis.
Learn tips from the field for managing the costs of antibodies for flow cytometry.
Bacteria such as E. coli are popular model systems for engineering and production of modified proteins. Yet the idea of putting bacteria in a cell sorter conjures unwelcome images. Take heart, the Sony SH800 cell sorter, simplifies decontamination by quickly and easily letting researchers replace key components that come in contact with the sample. Here are some examples, and publication citations.
Listen to Dr. Jennifer Doudna, one of the discoverers of CRISPR systems discusses how the new genome engineering technology was discovered. Video: Genome Engineering with CRISPR-Cas9: Birth of a Breakthrough Technology
Most commercially available antibodies contain small amounts of preservatives such as sodium azide to prevent microbial growth. However, sodium azide is also toxic to mammalian cells as it inhibits cellular respiration. Actual toxicity varies by cell type with neuronal cells being most sensitive. Toxicity is concentration, time, and temperature dependent. For most cell sorting experiments the health of cells are not impacted because the antibody is diluted and cells are typically incubated on ice for less than one hour.
Flow cytometry has long been considered a tool of hematologists and immunologists who primarily work with the hematopoetic system. Flow cytometry is capable of performing the simultaneous detection of more than 20 parameters; however it requires the sample to be prepared into a single cell suspension. The need for a single cell suspension makes the sample preparation more intensive for cells derived from tissues. Many tissues are also highly autofluorescent, which results in background noise that limits detection of low levels of expression on conventional flow cytometers.
With the ability to easily identify and isolate single cells from a heterogeneous phenotypic population, a cell sorter is a useful tool for laboratories studying and applying the CRISPR technology as a tool for editing genes and creating single cell libraries. CRISPR genome editing tools can be used to remove, change or insert DNA sequences into genes. The resulting mutations are maintained in the genome as cells proliferate –in contrast to transient transfection methods. Edits to genes are accomplished by directing the Cas9 nuclease to cleave DNA at a specific sequence and a guide RNA (gRNA) into the cell through transfection (or electroporation). For the edit to be completed the cell must be successfully transfected and the DNA cleaved by the Cas9 (referred to as cleavage efficiency). Many factors can impact cleavage efficiency including the efficiency of the transfection. A commonly used method to identify cells that have been successfully transfected is to include a plasmid encoding for a fluorescent protein such as eGFP in the transfection along with Cas9 and gRNA. Cells can then be sorted based on the expression of the fluorescent protein. The SH800 sorter, lets researchers deposit target cells into 96- or 384 multi-well devices and...
T-cells are part of the adaptive immune system with several subtypes including CD8+ effector T-cells and CD4+ helper T-cells. Main categories can be divided into several sub-categories. What are the critical markers to study T-cells? From the key subsets how can other subsets be defined?
Thank your super hero mom this Mother’s Day by giving her some quality time and something to remember—the gift of knowing about the work you do that makes the world a better place. To help you with what could be the best little talk ever here’s a 5-minute animation from the Royal Society that explains gene editing and the importance CRISPR-Cas-9 we hope you will find useful. Happy Mother’s Day to all, thank you for what you do. Video: What is gene editing and how does it work?