Enhanced detection of fluorescence quenching in labeled cells
Abstract
A method is provided for quantifying BrdU labeled DNA in cells. The BrdU is incorporated into the DNA and the DNA is stained with a first fluorochrome having a fluorescence which is quenchable by BrdU. The first fluorochrome is preferably a thymidine base halogen analogue, such as a Hoechst fluorochrome. The DNA is then stained with a second fluorochrome having a fluorescence that is substantially uneffected by BrdU. The second fluorochrome may be selected from the group consisting of mithramycin, chromomycin A3, olivomycin, propidium iodide and ethidium bromine. The fluorescence from the first and second fluorochromes is then measured to obtain first and second output signals, respectively. The first output signal is substracted from the second output signal to obtain a difference signal which is functionally related to the quantity of BrdU incorporated into DNA. The technique is particularly useful for quantifying the synthesis of DNA during the S-phase of the cell cycle.
- Inventors:
-
- Los Alamos, NM
- Issue Date:
- Research Org.:
- Los Alamos National Laboratory (LANL), Los Alamos, NM (United States)
- OSTI Identifier:
- 868152
- Patent Number(s):
- 5084378
- Assignee:
- United States of America as represented by United States (Washington, DC)
- Patent Classifications (CPCs):
-
C - CHEMISTRY C12 - BIOCHEMISTRY C12Q - MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS
Y - NEW / CROSS SECTIONAL TECHNOLOGIES Y10 - TECHNICAL SUBJECTS COVERED BY FORMER USPC Y10S - TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- DOE Contract Number:
- W-7405-ENG-36
- Resource Type:
- Patent
- Country of Publication:
- United States
- Language:
- English
- Subject:
- enhanced; detection; fluorescence; quenching; labeled; cells; method; provided; quantifying; brdu; dna; incorporated; stained; fluorochrome; quenchable; preferably; thymidine; base; halogen; analogue; hoechst; substantially; uneffected; selected; consisting; mithramycin; chromomycin; a3; olivomycin; propidium; iodide; ethidium; bromine; fluorochromes; measured; obtain; output; signals; respectively; signal; substracted; difference; functionally; related; quantity; technique; particularly; useful; synthesis; s-phase; cell; cycle; difference signal; labeled dna; functionally related; output signals; particularly useful; output signal; cell cycle; propidium iodide; /435/436/
Citation Formats
Crissman, Harry A, and Steinkamp, John A. Enhanced detection of fluorescence quenching in labeled cells. United States: N. p., 1992.
Web.
Crissman, Harry A, & Steinkamp, John A. Enhanced detection of fluorescence quenching in labeled cells. United States.
Crissman, Harry A, and Steinkamp, John A. Wed .
"Enhanced detection of fluorescence quenching in labeled cells". United States. https://www.osti.gov/servlets/purl/868152.
@article{osti_868152,
title = {Enhanced detection of fluorescence quenching in labeled cells},
author = {Crissman, Harry A and Steinkamp, John A},
abstractNote = {A method is provided for quantifying BrdU labeled DNA in cells. The BrdU is incorporated into the DNA and the DNA is stained with a first fluorochrome having a fluorescence which is quenchable by BrdU. The first fluorochrome is preferably a thymidine base halogen analogue, such as a Hoechst fluorochrome. The DNA is then stained with a second fluorochrome having a fluorescence that is substantially uneffected by BrdU. The second fluorochrome may be selected from the group consisting of mithramycin, chromomycin A3, olivomycin, propidium iodide and ethidium bromine. The fluorescence from the first and second fluorochromes is then measured to obtain first and second output signals, respectively. The first output signal is substracted from the second output signal to obtain a difference signal which is functionally related to the quantity of BrdU incorporated into DNA. The technique is particularly useful for quantifying the synthesis of DNA during the S-phase of the cell cycle.},
doi = {},
journal = {},
number = ,
volume = ,
place = {United States},
year = {Wed Jan 01 00:00:00 EST 1992},
month = {Wed Jan 01 00:00:00 EST 1992}
}
Works referenced in this record:
Flow cytometric analysis of factors which influence the BrdUrd-Hoechst quenching effect in cultivated human fibroblasts and lymphocytes
journal, January 1983
- Kubbies, M.; Rabinovitch, P. S.
- Cytometry, Vol. 3, Issue 4
Replication kinetics of Chinese hamster chromosomes as revealed by bivariate flow karyotyping
journal, January 1983
- Cremer, C.; Gray, J. W.
- Cytometry, Vol. 3, Issue 4
Dual-laser, differential fluorescence correction method for reducing cellular background autofluorescence
journal, November 1986
- Steinkamp, John A.; Stewart, Carleton C.
- Cytometry, Vol. 7, Issue 6, p. 566-574
Microfluorometric Detection of Deoxyribonucleic Acid Replication in Human Metaphase Chromosomes
journal, December 1973
- Latt, S. A.
- Proceedings of the National Academy of Sciences, Vol. 70, Issue 12