Flow Cytometry
Flow cytometry is a powerful technology for characterizing and analyzing cells. The fluidics, optics, and electronics systems in flow cytometry work together to simultaneously measure and analyze multiple physical characteristics of particles, usually cells, as they move in a fluid stream through a beam of light. A flow cytometer can tell us about a particle’s relative size, granularity or internal complexity, and fluorescence intensity. The Child & Family Research Institute (CFRI) Flow Core Facility provides equipment for flow cytometric analysis and full service cell sorting, as well as flow cytometry training and education for new users.

Analysis
The ability of flow cytometers to evaluate particles or cells at an extremely rapid rate (e.g. up to 25,000 events per second) makes this technology ideally suited for the reliable and accurate quantitative analysis of selected physical properties of the target particles or cells. Any suspended 0.2-50 micrometers particle or cell suspension of peripheral blood, bone marrow, body fluid or tissue can be labeled with multiple color antibody cocktails and run through the multi-laser flow cytometer for FACS analysis. Therefore, FACS has been widely used in applications such as: tracking GFP (YFP, CFP) expression; subset identification; cell activation status; cell cycle; surface protein expression; cytokine production; chemokine receptor repertoire; cell to cell interactions; cell signaling; cell viability; and apoptosis.          

The CFRI Flow Core Facility has the following equipment:

1. FACS Caliburs. There are two Caliburs, purchased in l998 and 2002 respectively. Both FACS Caliburs have highly sensitive blue (480 nm) and red (635 nm) lasers, which are used exclusively for quantitative analysis applications. The FACS Calibur is equipped with four fluorescence detectors to allow simultaneous analysis of six parameters (two light scatter and four fluorochromes). Data is acquired and analysed using CellQuestPro software. The FACS Calibur is engineered with fixed optical, electronic and fluidic components, giving it the flexibility for an investigator-operated instrument after basic training.
   
   
   
   
   
2. One BD FACSAria bought in 2005. The FACS Aria has blue (480 nm), red (635 nm) and violet (405 nm) lasers and is equipped with ten fluorescence detectors to allow simultaneous analysis of twelve parameters (two light scatter and ten fluorochromes) using FACS Diva software. The FACS Aria can be used for cell sorting and analyses.
   
   
   
   
   
3. One new BD LSRII was installed at the end of September 2009. The LSRII is equipped with four lasers – blue (480 nm), red (635 nm), green (532 nm) and violet (405 nm), and a HTS (High Throughput Sampler) device that allows automated acquisition of specimens in 96 well or 384-well plate format. It is capable of measuring twenty-one parameters using FACS Diva software.
   
   
   
   
   
4. A Biomek NX, liquid handler was purchased from Beckman Coulter in August 2009. Biomek NX sets the standard for flexible laboratory solutions. It puts every aspect of liquid handling – including pipetting, dilution, dispensing and integration – into a single, automated system that is as powerful as it is efficient and economical. For example, it can be used for preparation of FACS samples, cytokine detection assays, and automated PCR.
   
   
   
   
5. The flow core has two Mac computers running Flowjo software for FACS data analysis. In addition, some individual labs use Flowjo, FCS express or CellQuestPro software for their FACS data analyses.
   
   

Cell Sorting
One of the properties of flow cytometers is the ability to sort particles or cells by electronic deflection into a separate collection tubes, plates or slides. Because multiple fluorochromes can be assessed simultaneously by flow cytometry, it can easily separate rare events from complex mixtures of cells on the basis of multiple marker expression. Thus, flow cytometry is especially well suited for cell purification requiring high purity. The CFRI Flow Core Facility has a high-speed cell sorter, BD FACS Aria, which is capable of both analysis and cell sorting (e.g. up to ten distinct fluorescent probes reacting with different cell associated molecules). After sorting cells can be, for example, cultured, cloned, or processed for protein expression and function analysis or for mRNA analysis using quantitative PCR or gene array techniques, etc.

FACS Training
In addition to analysis and cell sorting, the CFRI Flow Core Facility offers comprehensive training on instruments (FACS Calibur, FACS Canto, LSRII and FACS Aria) and software (CellquestPro, FACS Diva, FlowJo and FCS Express). We can also consult on experimental design, fluocrome combinations and maximizing the effectiveness and cost of your experiments.

We aim to provide world-class flow cytometry services to CFRI students, staff, researchers and scientists. We also welcome outside users (please contact Lisa).    

Flow Core Committee:
Dr. Tobias Kollman (Current Chair); Dr. Rusung Tan (Current FACS Account Manager); Dr. Kirk Schultz (Previous FACS Account Manager); Dr. Jan Dutz; Dr. Megan Levings; Dr. Peter Van Den Elzen and Dr. Lixin (Lisa) Xu (Current Flow Core Manager).

Contact:
Dr. Lisa Xu. Tel. 604-875-2000 ext. 5987.

Last updated: 12/04/2009