We are working to develop flow-based methods for separating different cell types from a mixed-cell co-culture. We are currently pursuing two separate projects. In the first, we are co-culturing normal fibroblasts with transformed fibroblasts in order to determine if cell-cell interactions can overcome the growth inhibition which the normal cells experience in 3-D culture. In collaboration with Dr. Wright, we have developed subclones of each fibroblast cell type which are stably transfected with green fluorescent protein (GFP). We have characterized the stability of GFP expression in these cells using flow analysis, and are currently conducting experiments using simultaneous flow measurement of GFP and DNA content in order to measure the proliferative status of each cell type in a mixed-cell co-culture. The second project involves investigating the radiation response of monolayer and spheroid co-cultures of normal human prostate and fibroblast cells. We are inv esti gating several potential methods for using flow analysis and sorting to separate these cell types, including fuorescently-tagged antibodies directed against the cell surface, membrane labeling using PKH-dyes, and GFP transfection. New antibodies will be developed using phage display technology. Flow analysis will be critical in evaluating these methods and in isolating high-affinity antibodies for physical separation of the cells from a mixed-cell co-culture by the use of magnetic beads.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Biotechnology Resource Grants (P41)
Project #
5P41RR001315-17
Application #
6119968
Study Section
Project Start
1999-07-01
Project End
2000-06-30
Budget Start
1998-10-01
Budget End
1999-09-30
Support Year
17
Fiscal Year
1999
Total Cost
Indirect Cost
Name
Los Alamos National Lab
Department
Type
DUNS #
City
Los Alamos
State
NM
Country
United States
Zip Code
87545
Frumkin, Jesse P; Patra, Biranchi N; Sevold, Anthony et al. (2016) The interplay between chromosome stability and cell cycle control explored through gene-gene interaction and computational simulation. Nucleic Acids Res 44:8073-85
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Chaudhary, Anu; Ganguly, Kumkum; Cabantous, Stephanie et al. (2012) The Brucella TIR-like protein TcpB interacts with the death domain of MyD88. Biochem Biophys Res Commun 417:299-304
Marina, Oana C; Sanders, Claire K; Mourant, Judith R (2012) Correlating light scattering with internal cellular structures. Biomed Opt Express 3:296-312
Houston, Jessica P; Naivar, Mark A; Jenkins, Patrick et al. (2012) Capture of Fluorescence Decay Times by Flow Cytometry. Curr Protoc Cytom 59:1.25.1-1.25.21
Marina, Oana C; Sanders, Claire K; Mourant, Judith R (2012) Effects of acetic acid on light scattering from cells. J Biomed Opt 17:085002-1

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