Cyclins appear at specific points in the cell cycle. Cells undergo a protocol to permeabalize and fix the cell membrane in order to make intracellular components accessible. Intracellular cyclins are then labeled with monoclonal antibodies conjugated either directly or indirectly to fluorochromes.
Drs. Alex Nakeff and Fred Valeriote investigate the role of novel anticancer agents on perturbations of the cell cycle (see reference below). Intracellular anti-cyclin B1 antibody conjugated to fluorescein isothiocynate (FITC, y-axis) was counterstained with propidium iodide for DNA content (x-axis). Cyclin B1 expression is known to be maximal during mitosis. In the CellQuest* density plot above, the major population at the bottom left is G0- and/or G1-phase cells. S-phase cells appear with increasing DNA content and a slight but gradual increase in cyclin B1. G2-phase cells are located at the end of the S-phase trend; M-phase (motitic) cells have the same amount of DNA as G2-phase, but express higher levels of cyclin B1. (A second cycle with increasing DNA content can almost be seen as the cells undergo endoreduplication: the failure of cytokenesis cell division in mitosis, typically caused by improper spindle formation in eukaryotic cells.)
*CellQuest is a trademark of BD Biosciences.
Subramanian B, Nakeff A, Media J, Wentland M, Valeriote F. Cellular Drug Action Profile Paradigm Applied to XK469. J. Expt. Ther. Oncol. 2(5): 253-63, 2002.