Acridine orange accumulation in acid organelles of normal and vacuolated frog skeletal muscle fibres

Abstract

The spatial distribution of acid membrane organelles and their relationships with normal and vacuolated transverse tubules has been studied in living frog skeletal muscle fibres using confocal microscopy. Acridine orange (AO) was used to evaluate acid compartments, while a lipophilic styryl dye, RH 414, was employed to stain the membranes of the T-system. AO accumulated in numerous spherical granules located near the poles of nuclei and between myofibrils where they were arranged in short parallel rows, triplets or pairs. AO granules could be divided into three groups: green (monomeric AO), red (aggregated AO), and mixed green/red. As demonstrated by lambda-scanning, most granules were mixed. Double staining of muscle fibres with AO and RH 414 revealed almost all AO granules located near the transverse tubules. Vacuolation of the T-system was induced by glycerol loading and subsequent removal. The close juxtaposition of AO granules and the T-system was preserved in vacuolated fibres. The lumens of vacuoles did not accumulate AO. It is concluded that AO granules represent an accumulation of AO in lysosome-related organelles and fragmented Golgi apparatus and a possible functional role of the spatial distribution of such acidic compartments is discussed.

Real-time imaging of exocytotic mucin release and swelling in Calu-3 cells using acridine orange

“…AO is widely used to stain live cells where it accumulates at high concentrations in acidic intracellular compartments (typically pH 4.5–6.0) showing significant spectral shifts attributed to its aggregation [13–15]. To verify whether pH itself affects spectral properties of AO independently of its concentration we have measured emission spectra for the AO solutions of intermediate concentration of 4 mM and pH of 5.5 and 7.4, Fig.…”

“…AO is a widely used metachromatic stain for nucleic acids, when it intercalates with DNA it fluoresces green (525 nm), and when it interacts with RNA it fluoresces red (>630 nm) [12]. Such spectral shift is also observed when AO accumulates at high concentrations in acid intracellular compartments and is attributed to the formation of di- and oligomeric aggregates [13–15]. Although AO is one of the oldest fluorescent dyes, its properties at high concentrations favoring aggregated form are still largely unknown, including extinction coefficient or quantum yield.…”

Sequestration of AS-DACA into Acidic Compartments of the Membrane Trafficking System as a Mechanism of Drug Resistance in Rhabdomyosarcoma

“…This study only provided qualitative data on AS-DACA sequestration. However, to ensure that the endocytic inhibitors were exerting their desired effect on the endocytic pathway, acridine orange (AO) was included and its intracellular distribution observed by fluorescence [39]. AO was added at a final concentration of 2 μM for 20 min and was observed under wavelengths of 490 nm–500 nm and 520 nm to visualize accumulation of orange vesicles and nuclear distribution respectively.…”

“Turn off–on” phosphorescent biosensors for detection of DNA based on quantum dots/acridine orange

“…As a fluorescent pigment and an important nucleic acid probe, acridine orange (AO) is not only widely used to study the dynamics of DNA [28], but also commonly applied in chemistry and biology as a DNA intercalator, cell stain, and membrane penetrating probe [29][30][31][32].…”