Method for rapid isolation of sensitive mutants
Abstract
Sensitive mammalian cell mutants are rapidly isolated using flow cytometry. A first population of clonal spheroids is established to contain both normal and mutant cells. The population may be naturally occurring or may arise from mutagenized cells. The first population is then flow sorted by size to obtain a second population of clonal spheroids of a first uniform size. The second population is then exposed to a DNA-damaging agent that is being investigated. The exposed second population is placed in a growth medium to form a third population of clonal spheroids comprising spheroids of increased size from the mammalian cells that are resistant to the DNA-damaging agent and spheroids of substantially the first uniform size formed from the mammalian cells that are sensitive to the DNA-damaging agent. The third population is not flow sorted to differentiate the spheroids formed from resistant mammalian cells from spheroids formed from sensitive mammalian cells. The spheroids formed from sensitive mammalian cells are now treated to recover viable sensitive cells from which a sensitive cell line can be cloned.
- Inventors:
-
- Los Alamos, NM
- Issue Date:
- Research Org.:
- Los Alamos National Laboratory (LANL), Los Alamos, NM (United States)
- OSTI Identifier:
- 871073
- Patent Number(s):
- 5652098
- Assignee:
- United States of America as represented by United States (Washington, DC)
- Patent Classifications (CPCs):
-
G - PHYSICS G01 - MEASURING G01N - INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- DOE Contract Number:
- W-7405-ENG-36
- Resource Type:
- Patent
- Country of Publication:
- United States
- Language:
- English
- Subject:
- method; rapid; isolation; sensitive; mutants; mammalian; cell; rapidly; isolated; flow; cytometry; population; clonal; spheroids; established; contain; normal; mutant; cells; naturally; occurring; arise; mutagenized; sorted; size; obtain; uniform; exposed; dna-damaging; agent; investigated; placed; growth; medium; form; third; comprising; increased; resistant; substantially; formed; differentiate; treated; recover; viable; line; cloned; naturally occurring; flow cytometry; uniform size; mammalian cells; growth medium; cell line; mutant cells; flow sorted; /435/
Citation Formats
Freyer, James P. Method for rapid isolation of sensitive mutants. United States: N. p., 1997.
Web.
Freyer, James P. Method for rapid isolation of sensitive mutants. United States.
Freyer, James P. Wed .
"Method for rapid isolation of sensitive mutants". United States. https://www.osti.gov/servlets/purl/871073.
@article{osti_871073,
title = {Method for rapid isolation of sensitive mutants},
author = {Freyer, James P},
abstractNote = {Sensitive mammalian cell mutants are rapidly isolated using flow cytometry. A first population of clonal spheroids is established to contain both normal and mutant cells. The population may be naturally occurring or may arise from mutagenized cells. The first population is then flow sorted by size to obtain a second population of clonal spheroids of a first uniform size. The second population is then exposed to a DNA-damaging agent that is being investigated. The exposed second population is placed in a growth medium to form a third population of clonal spheroids comprising spheroids of increased size from the mammalian cells that are resistant to the DNA-damaging agent and spheroids of substantially the first uniform size formed from the mammalian cells that are sensitive to the DNA-damaging agent. The third population is not flow sorted to differentiate the spheroids formed from resistant mammalian cells from spheroids formed from sensitive mammalian cells. The spheroids formed from sensitive mammalian cells are now treated to recover viable sensitive cells from which a sensitive cell line can be cloned.},
doi = {},
journal = {},
number = ,
volume = ,
place = {United States},
year = {1997},
month = {1}
}
Works referenced in this record:
X-ray-sensitive mutants of Chinese hamster ovary cell line isolation and cross-sensitivity to other DNA-damaging agents
journal, December 1983
- Jeggo, P. A.; Kemp, L. M.
- Mutation Research/DNA Repair Reports, Vol. 112, Issue 6
Mutagen-sensitive cell lines are obtained with high frequency in V79 Chinese cells
journal, June 1987
- Zdzienicka, Małgorzata Z.
- Mutation Research/Fundamental and Molecular Mechanisms of Mutagenesis, Vol. 178, Issue 2
Viable sorting of intact multicellular spheroids by flow cytometry
journal, July 1987
- Freyer, James P.; Wilder, Mark E.; Jett, James H.
- Cytometry, Vol. 8, Issue 4
Use of xantham gum to suspend large particles during flow cytometric analysis and sorting
journal, November 1989
- Freyer, James P.; Fillak, David; Jett, James H.
- Cytometry, Vol. 10, Issue 6