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Enhanced detection of fluorescence quenching in labeled cells

Patent ·
OSTI ID:868152
 [1];  [1]
  1. Los Alamos, NM
+ Show Author Affiliations

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.

Research Organization:
Los Alamos National Laboratory (LANL), Los Alamos, NM
DOE Contract Number:
W-7405-ENG-36
Assignee:
United States of America as represented by United States (Washington, DC)
Patent Number(s):
US 5084378
OSTI ID:
868152
Country of Publication:
United States
Language:
English

References (4)

Dual-laser, differential fluorescence correction method for reducing cellular background autofluorescence journal November 1986
Flow cytometric analysis of factors which influence the BrdUrd-Hoechst quenching effect in cultivated human fibroblasts and lymphocytes journal January 1983
Microfluorometric Detection of Deoxyribonucleic Acid Replication in Human Metaphase Chromosomes journal December 1973
Replication kinetics of Chinese hamster chromosomes as revealed by bivariate flow karyotyping journal January 1983

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