Benzophenanthridine alkaloids as a new supravital DNA dyes.
| Authors | |
|---|---|
| Year of publication | 2006 |
| Type | Article in Proceedings |
| Conference | Sborník XIV. Cytoskeletálního klubu |
| MU Faculty or unit | |
| Citation | |
| Field | Genetics and molecular biology |
| Keywords | benzophenanthridine alkaloids; DNA dye; flow cytometry; fluorescence microscopy; multicolor analysis; cell cycle |
| Description | We have proved that macarpine (MA) and chelirubine (CHR) can be used as supravital fluorescent DNA probes both in fluorescence microscopy and flow cytometry. MA and CHR are quaternary benzo[c]phenanthridine alkaloids (QBA), that are naturally occurring products with noteworthy antiproliferative activity and fluorescence characteristics. QBA are commonly isolated from plants of Fumariaceae, Papaveraceae, Ranunculaceae and Rutaceae families. When added to intact cell suspensions, water stock solutions of the alkaloids (MA, CHR) stain the cells upon brief (in seconds) incubation at low final concentration (0,01-0,001 mg/ml). We have also evaluated the convenience of these minor QBA as supravital nucleic acid probes for time effective multiparameter immunophenotyping of peripheral blood and bone marrow leukocytes. Heparinized peripheral blood or bone marrow aspirate cells were stained on the surface with a combination of fluorochrome-conjuagted monoclonal antibodies directed against surface markers. Then, an appropriate amount of individual QBA was added and, upon dilution, the samples were acquired on a standard (FACSCalibur) or polychromatic (CyAn) flow cytometer. Our results show that spectral characteristics of individual QBA allow for simultaneous surface immunophenotyping and semiquantitative DNA determination. MA and CHR are excitable by common argon lasers (488 nm) emmiting at the range 575 - 750 nm (i.e. fluorescence channels 2-5). Fluorescence channels 1 and 5 – 7 are available for surface immunophenotyping on FACSCalibur and CyAn, respectively. |
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