Instrumentation
The optical specifications for each of the UC Davis Flow Cytometry Shared Resource’s cell sorters and cytometers are available online at the Fluorofinder website. Fluorofinder is free to join and houses a UC Davis Flow Cytometry Core page detailing the lasers and optical filters found on machines available at the shared resource.
Fluorofinder also offers a free multi-color panel building tool which harnesses the reagent catalogs of over 15 suppliers to help guide reagent selection and panel design. FCSR staff will operate the cell sorters to assist users in purifying specific cell subpopulations for their downstream applications. Staff are available to help with experimental design and will train investigators to independently operate the analytic flow cytometers.
Tupper Hall, UC Davis campus
Beckman Coulter “Cytoflex” Flow Cytometer: 4 laser (V,B,Y,R), 13-color
This Cytoflex analytical flow cytometer offers several new features for Davis campus users, including a plate-based sample loader and yellow laser for efficient red fluorescent protein detection. The Cytoflex is user-friendly and supports a wide range of projects, from single-color fluorescent protein screening to moderately complex immunophenotyping applications.
Becton Dickinson “LSR-II” Flow Cytometer: 3 laser (V,B,R), 16-color
This 3-laser analytical cytometer combines digital pulse processing and fast analog to digital converters to acquire up to 16 fluorescence signals per cell. High-powered violet (50mW 405nm), blue (100mW 488nm) and red (50mW 635nm) lasers excite a variety of fluorochromes. The violet laser excites dyes such as Pacific Blue, Brilliant Violet 421, DAPI, AmCyan, Pacific Orange, Brilliant Violet series dyes such as BV570, BV605, BV650, BV711 & BV785, in addition to several 'Quantum Dot' semi-conductor nanocrystal dyes.
The 488nm laser excites dyes such as FITC, green fluorescent protein, Alexa 488, PE, PE-Texas Red, PECy5, PECy5.5, PECy7. The red laser excites APC, Alexa 647, Alexa 700, and APCCy7. In 2017, a new HP computer was installed to accommodate a major upgrade in acquisition and analysis software, DIVA 8.0. These upgrades have streamlined maintenance and quality assurance and allow a large number of trained investigators to operate this cytometer independently.
Cytek “Aurora” Flow Cytometer: 5 laser (UV,V,B,Y,R), 40+ color
The Cytek Aurora is a 5-laser, 40+ color analytic cytometer that was recently added to the core. This cytometer uses 5 cutting-edge spectral array multi-window photodetectors to capture the spectral characteristics of the dyes attached to each sample, and then uses advanced spectral deconvolution software to unmix these signals to achieve unprecedented multiplexing capability. Currently, up to 40 colors can be detected using 5 lasers (355, 405, 488, 561 and 640nm). Several dyes that would be indistinguishable using traditionally configured cytometers are now able to be used simultaneously to discriminate biomarker expression, thus increasing the number of useful fluorochromes to 40 or more per panel.
The Astrios EQ high-speed cell sorter is a sophisticated and flexible cell sorting system. The Astrios is currently equipped with 4 lasers, including 405nm violet, 488 nm blue, 561 nm yellow and 640 nm red. In this set-up this instrument is capable of the simultaneous detection of up to 18 colors and two scatter parameters. The fluorescence collection optics allow for the future addition of up to 3 other lasers as research needs evolve. The Astrios offers unparalleled operator and sample protection from biohazards; its sample injection and cell deposition systems are fully enclosed in a Baker Class II biological safety cabinet and continuously exhausted to the HEPA filter in the BSCs hood. Up to six populations of cells can be sorted simultaneously and reliably, with the option to select individual sort stringencies per stream. Other features of the Astrios EQ sorter include the following:
- 6, 4 and 2 way sorting into tubes, 96 & 384wp deposition, sample tube format flexibility.
- Jet in Air design: avoids damaging acceleration effects of cuvette sorter.
- Nozzle sizes from 50-200um, stable low pressure regulation for 200um large cell sorting.
- Ease of use features: Automated laser and fluidics startup, simplified laser alignment, automated QC routines, beadless drop delay (standard bead-based optional), automated droplet breakoff and side stream monitoring during sorting, post-clog sample and product protection.
- Fast event-processing electronics afford high speed sample enrichment rates of up to 70,000 events per second and cell purification rates of up to 25,000 cells per second for well-dispersed samples.
Fluidigm BioMark HD
Many investigators require single cell gene analysis for their research. The Fluidigm BioMark microfluidics based single cell nucleic acid analysis platform has been placed in the FCSR to allow users to proceed directly from live cell sorting experiments to genetic analysis. This system is designed to perform single cell expression analysis, expression analysis from limited RNA samples, digital PCR/copy number variation, haplotyping, genotyping, SNP mutation detection, and microarray validation.
College of Biological Sciences Satellite Facility, Davis Campus
Becton Dickinson “FACS Canto II” Flow Cytometer: 2 laser (B,R), 6-color
This 6-color analyzer provides local access to basic analytic cytometry equipment for investigators in the College of Biological Sciences in Briggs Hall. The Canto II was donated at no cost to the FCSR in 2015 through collaboration with the Howard Hughes Medical Institute in Seattle, WA. UC Davis department chairs Drs. Wolf-Dietrich Heyer and Jodi Nunnari provided funds to relocate and refurbish the cytometer and renovated laboratory space to house it. The Canto II is now operated by the FCSR for recharge-based usage and is open to use to all campus departments.
UC Davis Medical Center, Sacramento Campus
Becton Dickinson “Fortessa” Flow Cytometer: 5-laser (UV,V,B,Y,R), 18-color
The Fortessa is an advanced analytical cytometer that is configured with 5 high-powered lasers and multiple photodetector arrays to collect fluorescence from a wide variety of fluorochromes. This cytometer is equipped with 60mW UV (350nm), 100mW violet (405nm), 100mW blue (488nm), 100mW yellow (561nm) and 100mW red (640nm) lasers, allowing for the simultaneous measurement of up to 18 fluorescent readouts and two scatter signals.
The UV laser provides the low wavelength excitation necessary for calcium flux assays (Indo-1 blue and violet), DAPI excitation for DNA content analysis and the newer Brilliant Ultra Violet dyes such as BUV 395 and 737. Signals from violet laser (405nm) excitation terminate in an array of six photodetectors that can measure a variety of Brilliant Violet dyes, including BV421, 570, 605, 650, 711 and 786, as well as other traditional fluorochromes such as Pacific Blue, Aqua-emitting Live/Dead stain, DAPI and several Quantum Dot dyes. The 488nm laser is used for GFP, FITC, Alexa488 and PerCPCy5.5 or propidium iodide. The 100mW 561nm yellow-green laser efficiently excites phycoerytherin (PE) and PE tandem dyes such as PE-Texas Red, PE-Cy5, PE-Cy5.5 and PE-Cy7. A highly powered red laser improves the excitation of dyes such as APC, Alexa 647, Alexa 700 and APC-Cy7 (and its analogs). A high throughput sampling module increases sample loading and acquisition rates.
Becton Dickinson “InFlux” Cell Sorter: 5-laser (UV,V,B,Y,R), 16-color
This high-speed 5-laser cell sorter is located in Sacramento to provide operator-assisted cell sorting to researchers in Sacramento and from the Davis campus. The inFlux sorter is equipped with five high-powered lasers to provide 100mW UV (355nm), 50mW violet (405nm), 200mW blue (488nm), 150mW green (532nm) and 50mW red (635nm) excitation beams and is configured to detect two-light scatter and 16 individual fluorescent markers. Recent upgrades include an Accudrop automated drop delay calibration system that will streamline cytometer setup prior to sorting and an advanced software package that offers improved sort setup and post-sort analysis tools.
Becton Dickinson “Aria II” Cell Sorter: 3-laser (V,B,R), 8-color in HEPA filtered enclosure
The BD FACS Aria II cell sorter is available for high-speed cell sorting in the Sacramento flow cytometry facility. The Aria II is capable of detecting up to 8 colors simultaneously and offers excellent fluorescent resolution. The Aria II takes advantage of the powerful DIVA acquisition and cell sorting software and can be configured with 70, 85 and 100um nozzles to accommodate various cell types. The Aria is available for staff-assisted cell sorting and for independent use by approved investigators.