Staining of mitochondrial membranes with 10-nonyl acridine orange, MitoFluor Green, and MitoTracker Green is affected by mitochondrial membrane potential altering drugs.

Background: We set out to develop an assay for the simultaneous analysis of mitochondrial membrane potential and mass using the probes 10-nonyl acridine orange (NAO), MitoFluor Green (MFG), and MitoTracker Green (MTG) in HL60 cells. However, in experiments in which NAO and MFG were combined with orange emitting mitochondrial membrane potential (DeltaPsi(m)) probes, we found clear responses to DeltaPsi(m) altering drugs for both probes.

Methods: The three probes were titrated to determine whether saturation played a role in the response to drugs.

Simultaneous analysis of relative DNA and glutathione content in viable cells by phase-resolved flow cytometry.

Background: Analysis of the DNA cell cycle and glutathione content cannot be performed on viable cells, because the fluorescence emissions of the DNA-specific probe Hoechst 33342 and the glutathione-specific probe monobromobimane overlap completely. We decided to explore whether the emissions could be resolved by the singlet excited state lifetimes of the probes.

Methods: Viable cells were first incubated with Hoechst 33342 at 37 degrees C for 30 min and then with monobromobimane at room temperature for 10 min.

The mouse Y chromosome: enrichment, sizing, and cloning by bivariate flow cytometry.

In this report, we demonstrate the utility of interleukin-2 (IL-2) stimulation of spleen cell cultures and bivariate flow cytometry in the analysis and purification of the C57BL/6J mouse Y Chromosome. We determined that the DNA content of the C57BL/6J Y Chromosome is approximately 94.7 Mb, making it similar in size to human Chromosome 16 and significantly larger than previous estimates.

New flow cytometric technologies for the 21st century.

The envelope that defines the limits within which flow cytometry was developed is being rapidly expanded. For example: detection sensitivity has been extended to single molecules, the size range of "particle" analysis now extends from DNA fragments to plankton (1,000.+ microns), cell and chromosome sorting rates are being increased dramatically by using inactivation procedures (50,000 per second versus 2,000 per second), rapid kinetic flow cytometry enables real-time analysis of molecular assembly and cell function in the sub-second time domain, the lifetime of a fluorochrome bound to a single cell can be measured with nsec precision, and classical karyotype information (cell to cell heterogeneity) can be determined in a flow based system.