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UNM Shared Flow Cytometry

MSC 09 5025
1 University of New Mexico
Albuquerque, NM 87131

Physical Location:
Cancer Research Facility (CRF; building 229)
Room 217

Phone: (505) 272-0835

UNM Shared Flow Cytometry and High Throughput Screening Resource

HyperCyt®

The HyperCyt®high-throughput flow cytometry platform interfaces a flow cytometer and autosampler. As the sampling probe of the autosampler moves from one well to the next of a multi-well microplate, a peristaltic pump sequentially aspirates sample particle suspensions from each well. Between wells, the continuously running pump draws a bubble of air into the sample line. This results in the generation of a tandem series of bubble-separated samples for delivery to the flow cytometer. Sample and bubble volumes are determined by the time that the autosampler probe is in a microplate well or above a well intaking air. Accurate quantitative measurements have been demonstrated in endpoint assays at rates of 20 to 40 samples/min over a 4-decade range of fluorescence intensity using input cell concentrations of 1–20 million/mL and source well volumes of 5-15 uL. Typical sample volumes of 1-2 uL allow scarce quantities of test cells or reagents to be efficiently used.

As the air bubble-separated samples are delivered in a continuous stream to the flow cytometer, the data are likewise collected in a continuous stream, the accumulated data from all wells of a microplate representing a single data file. The time-resolved data, with periodic gaps corresponding to the passage of the sample-separating air bubbles, are analyzed by proprietary software developed at UNM. HyperCyt®technology has been licensed by IntelliCyt Corporation (http://www.intellicyt.com) and a commercial platform is currently available that is compatible with most commercial flow cytometry instruments.

In 2005, the New Mexico Molecular Libraries Screening Center, specializing in high throughput flow cytometry screening with the HyperCyt®platform, was selected as 1 of 10 institutions in the USA to participate in the pilot phase of the NIH-sponsored Molecular Libraries Screening Network, an NIH Roadmap initiative. The NMMLSC has since been renamed theUniversity of New Mexico Center for Molecular Discovery (UNMCMD)and selected in 2009 to participate in the production phase of the NIH screening initiative, the Molecular Libraries Probe Production Centers Network (MLPCN). The goal of this national network is to produce innovative chemical probes for use in biological research and drug development.

To date descriptions and results of more than 100 HyperCyt®screening projects have been published on the publicly accessible PubChem website (http://www.ncbi.nlm.nih.gov/sites/entrez?db=pcassay). These results represent a total of more than 4 million quantitative fluorescence measurements of biological interactions with small molecule compounds from the NIH Small Molecule Repository.

HyperCyt Drawing

HyperCyt in Action

Example of HyperCyt time-resolved Data

HyperCyt®Literature

Peer Reviewed Publications

  1. Kuckuck, F. W., Edwards, B. S., and Sklar, L. A. High throughput flow cytometry. Cytometry,44:83-90, 2001.
  2. Jackson, W. C., Bennett, T. A., Edwards, B. S., Prossnitz, E., Lopez, G. P., and Sklar, L. A. Performance of in-line microfluidic mixers in laminar flow for high-throughput flow cytometry. Biotechniques,33:220-226, 2002.
  3. Jackson, W. C., Kuckuck, F., Edwards, B. S., Mammoli, A., Gallegos, C. M., Lopez, G. P., Buranda, T., and Sklar, L. A. Mixing small volumes for continuous high-throughput flow cytometry: Performance of a mixing Y and peristaltic sample delivery. Cytometry,47:183-191, 2002.
  4. Ramirez, S., Aiken, C. T., Andrzejewski, B., Sklar, L. A., and Edwards, B. S. High-throughput flow cytometry: Validation in microvolume bioassays. Cytometry,53A:55-65, 2003.
  5. Young, S. M., Curry, M. S., Ransom, J. T., Ballesteros, J. A., Prossnitz, E. R., Sklar, L. A., and Edwards, B. S. High-throughput microfluidic mixing and multiparametric cell sorting for bioactive compound screening. J Biomol Screen,9:103-111, 2004.
  6. Bartsch, J. W., Tran, H. D., Waller, A., Mammoli, A. A., Buranda, T., Sklar, L. A., and Edwards, B. S. An investigation of liquid carryover and sample residual for a high-throughput flow cytometer sample delivery system. Anal Chem,76:3810-3817, 2004.
  7. Young SM, Bologa C, Prossnitz ER, Oprea TI, Sklar LA, Edwards BS. High-throughput screening with HyperCyt flow cytometry to detect small molecule formylpeptide receptor ligands. J Biomol Screen 2005;10(4):374-82.
  8. Edwards BS, Bologa C, Young SM, Balakin KV, Prossnitz ER, Savchuck NP, Sklar LA, Oprea TI. Integration of virtual screening with high-throughput flow cytometry to identify novel small molecule formylpeptide receptor antagonists. Mol Pharmacol 2005;68(5):1301-10.
  9. Edwards BS, Young SM, Oprea TI, Bologa CG, Prossnitz ER, Sklar LA. Biomolecular screening of formylpeptide receptor ligands with a sensitive, quantitative, high-throughput flow cytometry platform. Nat Protoc 2006;1(1):59-66.
  10. Bologa CG, Revankar CM, Young SM, Edwards BS, Arterburn JB, Kiselyov AS, Parker MA, Tkachenko SE, Savchuck NP, Sklar LA, Oprea TI, Prossnitz ER. Virtual and biomolecular screening converge on a selective agonist for GPR30. Nat Chem Biol 2006;2(4):207-12.
  11. Saunders MJ, Kim H, Woods TA, Nolan JP, Sklar LA, Edwards BS, Graves SW. Microsphere-based protease assays and screening application for lethal factor and factor Xa. Cytometry A 2006;69(5):342-52.
  12. Edwards BS, Ivnitski-Steele I, Young SM, Salas VM, Sklar LA. High-throughput cytotoxicity screening by propidium iodide staining. Curr Protoc Cytom 2007;Chapter 9:Unit9 24.
  13. Simons PC, Young SM, Gibaja V, Lee WC, Josiah S, Edwards BS, Sklar LA. Duplexed, bead-based competitive assay for inhibitors of protein kinases. Cytometry A 2007;71(7):451-9.
  14. Dennis MK, Bowles HJ, MacKenzie DA, Burchiel SW, Edwards BS, Sklar LA, Prossnitz ER, Thompson TA. A multifunctional androgen receptor screening assay using the high-throughput Hypercyt flow cytometry system. Cytometry A 2008;73(5):390-9.
  15. Ivnitski-Steele I, Larson RS, Lovato DM, Khawaja HM, Winter SS, Oprea TI, Sklar LA, Edwards BS. High-throughput flow cytometry to detect selective inhibitors of ABCB1, ABCC1, and ABCG2 transporters. Assay Drug Dev Technol 2008;6(2):263-76.
  16. Winter SS, Lovato DM, Khawaja HM, Edwards BS, Steele ID, Young SM, Oprea TI, Sklar LA, Larson RS. High-throughput screening for daunorubicin-mediated drug resistance identifies mometasone furoate as a novel ABCB1-reversal agent. J Biomol Screen 2008;13(3):185-93.
  17. Young SM, Bologa CM, Fara D, Bryant BK, Strouse JJ, Arterburn JB, Ye RD, Oprea TI, Prossnitz ER, Sklar LA, Edwards BS. Duplex high-throughput flow cytometry screen identifies two novel formylpeptide receptor family probes. Cytometry A 2009;75(3):253-63.
  18. Edwards BS, Young SM, Ivnitsky-Steele I, Ye RD, Prossnitz ER, Sklar LA. High-content screening: flow cytometry analysis. Methods Mol Biol. 2009;486:151-65.
  19. Haynes MK, Strouse JJ, Waller A, Leitao A, Curpan RF, Bologa C, Oprea TI, Prossnitz ER, Edwards BS, Sklar LA, Thompson TA. Detection of intracellular granularity induction in prostate cancer cell lines by small molecules using the HyperCyt high throughput flow cytometry system. J Biomol Screen 14:596-609, 2009.

Reviews

  1. Waller, A., Simons, P., Prossnitz, E. R., Edwards, B. S., and Sklar, L. A. High throughput screening of G-protein coupled receptors via flow cytometry. Comb Chem High Throughput Screen,6:389-397, 2003.
  2. Edwards, B. S., Oprea, T. I., Prossnitz, E. R., and Sklar, L. A. Flow cytometry for high throughput, high content screening. Curr Opin Chem Biol,8:392-396, 2004.
  3. Waller, A., Simons, P. C., Biggs, S. M., Edwards, B. S., Prossnitz, E. R., and Sklar, L. A. Techniques: GPCR assembly, pharmacology and screening by flow cytometry. Trends Pharmacol Sci,25:663-669, 2004.
  4. Edwards, B,S., Sklar, L.A. Automation and high throughput flow cytometry. In: Flow Cytometry and Biotechnology, L.A. Sklar, Ed., Oxford University Press Inc., New York, NY, pp. 40-56, 2005.
  5. Edwards BS, Young SM, Saunders MJ, Bologa C, Oprea TI, Ye RD, Prossnitz ER, Graves SW, Sklar LA: High Throughput Flow Cytometry for Drug Discovery. Expert Opin Drug Discovery 2:685-696, 2007.
  6. Sklar LA, Carter MB, Edwards BS. Flow cytometry for drug discovery, receptor pharmacology and high-throughput screening. Curr Opin Pharmacol 2007;7(5):527-34.
  7. Salas VM, Edwards BS, Sklar LA. Advances in multiple analyte profiling. Adv Clin Chem 2008;45:47-74.