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Research

Flow Cytometry

URMC / Research / Flow Cytometry / Library

 

Library

The following reference materials are available for researchers to borrow:

  • Methods in Cell Biology, Volume 75: Cytometry 4th Edition. New Developments. (2004) Darzynkiewick, Roederer and Tanke (eds)
  • Current Protocols in Flow Cytometry
  • Course Notebook - 27th Annual Course in Flow Cytometry
    PROTOCOLS

Frequently Asked Questions:

This page is a work in progress and will be updated regularly.  If you have suggestions please let us know.

Updated as of Nov. 2020 - Here

Connecting to Smartsheet

All University employees have free Collaborator access to Smartsheet and account set up is done using your Active Directory credentials.  Although it is not required to have a Smartsheet account to access and submit a sort form, it is recommended as you will need an account to view or edit forms submitted via Smartsheet.  

Connect to Smartsheet in order to view or your edit sort forms.  For help with setting up your Smartsheet access click here.

Accessing the Smartsheet Sort Forms

The instrument specific sort forms can be found here - Statler, Waldorf, Agnes Stonewick and Link Hogthrob. 

For help filling out sort forms click here.

Software Guides

Software guides are available in the "User Docs" folder located on every instrument computer as well as in the FCC_Library (links below) server folder.

Troubleshooting and After Hours

Troubleshooting and after hours guides are available in the "User Docs" folder located on every instrument computer as well as in the FCC_Library (links below) server folder.

Peak 6 Bead Information

The reference bead that is used in the facility is from Spherotech. We use the Rainbow Calibration Particles, 6th peak of 8. The part number for this product is RCP-30-5A-6 and the current lot that we use is AK01.

Connecting to the FCC_Transfer server

At the instrument computer you can click the "shortcut to transfer" icon on the desktop of the instrument computer. An explorer window with a folder for each PI will open. NOTE - you will be able to view the contents of the folders but will not be able to read the contents of any of these folders, even your own. These folders are write only from the instruments. As before, drag the folders you want to transfer from the FCS folder to your group folder.  Also please note that this is a transfer space and not for long term storage.  Data will be removed regularly.

To recover your data on your lab computer:

There are 4 important points for accessing the pipeline from your lab computer:

  1. You must be connected to the URMC intranet.  You can not be on the guest network or River Campus network.
  2. You must use your URMC credentials.  Same as to access your URMC email.
  3. You must be using a University managed computer.  It does not work on a personal computer even if its capable of using VPN to access the intranet.
  4. Always use the full file path including the PI folder name.  You can not access the general pipeline folder and then click through to your specific folder.

File Path for PC:

Windows 7:

Click "Start" then in the "Search programs and files" field type:

Medical Center: \\smdnas\FCC_Transfer\PILastName_FirstInitial (ie. bushnell_t)

River Campus: \\smdnas.urmc-sh.rochester.edu\FCC_Transfer\PILastName_FirstInitial (ie. bushnell_t)

 

File Path for Mac:

Click "Go" in Finder, choose "Connect to Server" then type:

Medical Center: smb://smdnas/fcc_transfer/PILastName_FirstInitial (ie. bushnell_t)

River Campus: smb://smdnas.urmc-sh.rochester.edu/fcc_transfer/PILastName_FirstInitial (ie. bushnell_t)


If you are prompted for a username and password use your URMC
network credentials (what you would use for webmail or to log onto a
Windows computer here).

REMEMBER!!

Data will be automatically removed from the transfer partition after two weeks.

Connecting to the FCC_Library server

This is done similarly to the Transfer space. Here we have shared documents that are available to all registered users. This includes past lectures, IDEAS software, and other goodies.

For PC:

Start Menu >In the search menu type:

\\smdnas\fcc_library

For Mac:

Finder >Go > Connect to server

smb://smdnas/fcc_library

If you are prompted for a username and password use your URMC
network credentials (what you would use for webmail or to log onto a
Windows computer here).

Current Instrument Baseline

Available upon request

Grant Information and publication acknowledgements

Acknowledgements are always appreciated and are an important metric for the FCR in measuring our success.  For more information on this please visit the main CART website here.

The full summary appendix for all shared resource laboratories and be found on the homepage of the main CART website and a section of that document is included below.

The URMC Flow Cytometry Resource (FCR), under the scientific direction of Timothy Bushnell, Ph.D. and the technical direction of Matt Cochran, MS., is a full service facility with approximately 1100 sq. feet of newly renovated lab space, employing a staff of 9. The Core currently has traditional analytical tools including a BD Accuri C6+ (4-colors), 2 LSR-Fortessas (5 lasers and 18 colors) and 2 Symphony A1s (4 lasers and 16 colors) from BD Biosciences. Additional analytical instruments include 2 Cytek Aurora full spectrum cytometers. We also have 3 cell sorters, two BD FACSAria-II and a Cytek Aurora CS for sorting projects. In addition we have several non traditional analytical cytometers. The Amnis ImageStream MarkII allows for imaging flow cytometry and the detection of up to 10 fluorescent parameters with cellular localization information via its imaging capabilities. The Helios Mass Cytometer, which uses lanthanide metal mass spectrometry as opposed to traditional fluorescent tags, allowing the analysis of upwards of 50 parameters to be studied simultaneously. The resource also added the Nanosight NS3000 in 2015 for the detection of nano sized particles. The Nanosight is capable of measuring the size distribution and concentration of particles as small as 10 nanometers by using light scatter and Brownian motion. In July 2022 the resource also added a BioRad BioPlex200 for multiplex bead array assays measuring a wide range of biomarkers in solution. Two other instruments available in the resource are the Celigo, a microwell plate-based imaging cytometer, and the Seahorse XFe96 for measuring metabolic function.  Instrumentation is only part of what we offer. We also offer comprehensive training for staff, consultation for maximizing the effectiveness and cost of your experiments, as well as an environment that fosters success in both existing methods and crossing the boundaries into new frontiers.  All data generated in the FCC is archived onto a 10-terabyte server with daily tape backup.

The FCC has established a series robust QA/QC protocols and procedures that are designed to ensure the instruments are performing optimally. These include standard quality control measurements, as recommended by the instrument manufacturers. Additional detailed instrumentation characterization and QC are performed based on best practices in the field. All QA/QC information is available to investigators and instrumentation characterization data is available on the FCC’s website.

All investigators go through a robust training program to allow them run the analyzers independently, allowing for experienced users to have 24/7 access. All booking is done via an on-line calendar system. All sorting is done by the FCC staff, and is available to investigators during normal business hours and outside that on some occasions.

Recent Publications

  1. Enos, M., Bancos, S., Bushnell, T., and Crispe, I.N. (2008) E2F4 modulates differentiation and gene expression in hematopoietic progenitor cells during commitment to the lymphoid lineage J Immunol 180:3699-707
  2. McGrath, K.E., Kingsley, P.D., Koniski, A..D., Porter, R.L., Bushnell, T.P., and Palis, J.(2007) Enucleation of primitive erythroid cells generates a transient population of "pyrenocytes" in the mammalian fetus. Blood 111:2409-17
  3. Guzman, M.L., Li, X., Corbett, C.A., Rossi, R.M., Bushnell, T., Liesveld J.L., Herbet, J., Young, F., and Jordan C.T. (2007) Rapid and selective death of leukemia stem and progenitor cells induced by the compound 4-benzyl, 2-methyl, 1,2,4-thiadiazolidine, 3,5 dione (TDZD-8). Blood 110:4436-44
  4. Ryan, E.P., Bushnell, T.P., Friedman, A.E., Rahman, I., and Phipps, R.P. (2007) Cyclooxygenase-2 selective inhibition reduces B lymphoma survival by inducing oxidative stress and decreasing intracellular glutathione Cancer Immunol Immunother  57:347-58
  5. Neering, S.J., Bushnell, T., Sozer, S., Ashton, J., Rossi, R.M., Wang, P.-Y., Bell, D.R., Heinrich, D., Bottaro, A, and Jordan C.T. (2007) Leukemia stem cells in a genetically defined murine model of blast crisis CML. Blood 110:2578-85.
  6. Graf, B., Bushnell, T., and Miller , J. (2007) LFA-1-mediated T cell costimulation through increased localization of TCR/class II complexes to the cSMAC and exclusion of CD45 from the immunological synapse. J Immunol., 179:1616-24
  7. Neff-LaFord, H., Teske, S., Bushnell, T.P., and Lawrence, B.P. (2007) Aryl hydrocarbon receptor activation during influenza virus infection unveils a novel pathway of IFN-a production by phagocytic cells J Immunol 179:247-55
  8. Kingsley, PD, Malik J, Emerson RL, Bushnell TP, McGrath KE, Bloedorn LA, Bulger M, and Palis J (2006) “Maturational” globin switching in primary primitive erythroid cells.  Blood 107:1665-1672