Cell experiment:

Yeast cells in log phase are fixed in cold ethanol (70% by vol.) and stored at -20℃. Fixed cells are washed three times with cold 10 mM Tris/HCI pH 7, then mildly sonicated to eliminate aggregates and finally counted. Yeast cells are added to 45 μM Acridine Orange 10-Nonyl Bromide and incubated for 15 min at 20℃. Cells are centrifuged (3000×g, 5 min) then washed twice in 10 mM Tris/HCl pH 7. Red fluorescence of Acridine Orange 10-Nonyl Bromide bound to 106 yeast cells is measured at 640 nm and correlated to the calibration curve run with thin-walled vesicles containing known amounts of cardiolipin[2].

Acridine Orange 10-Nonyl Bromide is a fluorescent probe for cardiolipin (λex: 489 nm, λem: 525 nm).

Acridine Orange 10-Nonyl Bromide is a fluorescent probe for cardiolipin (λex: 489 nm, λem: 525 nm) which can be used to quantify the cardiolipin in isolated mitochondria[1]. when Acridine Orange 10-Nonyl Bromide interacts with cardiolipin, the dye excitation and emission wave lengths shift from 496 and 525 nm to 450 and 640 nm, respectively. Increasing amounts of cardiolipin (0 to 30 μM) and other acidic phospholipids in thin-walled vesicles added to Acridine Orange 10-Nonyl Bromide (45 μM) changes the red fluorescence emission measured at 640 nm according to the liposome composition[2].

[1]. Ratinaud MH, et al. In situ flow cytometric analysis of nonyl acridine orange-stained mitochondria from splenocytes. Cytometry. 1988 May;9(3):206-12. [2]. Gallet PF, et al. Direct cardiolipin assay in yeast using the red fluorescence emission of 10-N-nonyl acridine orange. Eur J Biochem. 1995 Feb 15;228(1):113-9.