Mechanism of radiosensitization by halogenated pyrimidines: effects of BrdU
on radiation induction of DNA and chromosome damage and its correlation with cell killing.
The specific primer sequences were obtained from Sigma Chemicals, USA; proprep protein extraction kit from InTron Biotechnology, USA, and Brdu
labeling kit from Roche, Germany.
incorporation, an assay for active cell proliferation, showed that incubation with 150 nm triptolide for 24h resulted in significantly fewer BrdU
-positive cells than incubation without triptolide (Fig.
Percentages of BrdU
positives and apoptotic cells were determined separately in each cellular compartment (parenchyma and stroma) of the whole mammary section (at least 2,000 epithelial cells and 1,000 stromal cells per section).
assay inhibition of proliferation was observed at > 20 [micro]M for NHDF and > 150 [micro]M for HaCaT keratinocytes.
The percentage of cells with 2N DNA that did not incorporate BrdU
DNA synthesis was assessed by a BrdU
ELISA kit according to the manufacturer's instruction.
Cytotoxicity of the compounds to P4CCR5 cells was determined in parallel using the Brdu
Cell Proliferation ELISA with chemilu-minescent detection (Roche Diagnostics GmbH, Mannheim, Germany) as previously described (Wagner et al.
Additional sections were taken for determination of cell labeling indices (percentages of cells in S phase) by BRDU
incorporation using the method of Soames et al.
The immunohistochemistry of hippocampus BrdU
positive cells had returned to the normal level in the group of depressive rats with low dosage Rhodiola rosea extract.
We purchased BrdU
and Hoechst 33258 from Eastman Kodak (Rochester, NY, USA), and Giemsa from Merck (Darmstadt, Germany).
l mM BrdU
was included in the medium for the first 48 h of culture in order to inhibit fibroblast growth.