Acridine Orange hydrochloride

Acridine orange is a cell-permeable fluorescent dye that binds to nucleic acids, resulting in an altered spectral emission.

In Vitro:Acridine orange has been employed extensively as a cytochemical stain and has shown to stain differentially DNA and RNA, and double-restranded nucleic acids in situ. Acridine orange can either intercalate into double helical nucleic acids (green fluorescence at 530 nm), or bind electrostatically to phosphate groups of single-stranded molecules (red fluorescence at 640 nm). This unique characteristic makes acridine orange useful for cell-cycle studies[1]. Acridine orange staining of unfixed cells may be used as a simple, fast means of obtaining information on cell ploidy levels and cell cycle status from DNA measurements (green fluorescence), and cell transcriptional activity from RNA staining (red fluorescence), in human and murine cells lines, peripheral blood and bone marrow specimens from patients with leukemia and mitogenically (phytohemagglutinin) or antigenically (mixed lymphocyte culture) stimulated human peripheral blood cultures[2].

References:
[1]. McMaster GK, et al. Analysis of single- and double-stranded nucleic acids on polyacrylamide and agarosegels by using glyoxal and acridine orange. Proc Natl Acad Sci U S A. 1977 Nov;74(11):4835-8. [2]. Traganos F, et al. Simultaneous staining of ribonucleic and deoxyribonucleic acids in unfixed cells using acridine orange in a flow cytofluorometric system. J Histochem Cytochem. 1977 Jan;25(1):46-56.

Chemoresistance to concanamycin A1 in human oral squamous cell carcinoma is attenuated by an HDAC inhibitor partly via suppression of Bcl-2 expression.[Pubmed:24278362]

PLoS One. 2013 Nov 20;8(11):e80998.

V-ATPase is involved in the acidification of the microenvironment around/in solid tumors, such as oral squamous cell carcinoma (OSCC). V-ATPase is thought to induce tumor invasion and multi-drug resistance in several malignant tumors, and it also contributes to maintaining the intracellular pH under an acidic microenvironment by inducing proton extrusion into the extracellular medium. However, there is little information regarding the effects of V-ATPase inhibitors on OSCCs. In this study, the effects of a V-ATPase inhibitor, concanamycin A1 (CMA), on the proliferation and apoptosis of OSCC were investigated in vitro. We used four OSCC cell lines, MISK81-5, SAS, HSC-4 and SQUU-B. Acridine orange staining revealed that the red fluorescence was reduced in all of the low concentration CMA-treated OSCC cells, indicating that the acidification of vesicular organelles in the OSCCs was prevented by the treatment with low-concentration of CMA. CMA treatment induced apoptosis in MISK81-5, SAS and HSC-4 cells, but not in SQUU-B cells. The p-p38 expression was not altered in CMA-treated SQUU-B cells, but their levels were increased in the other cells. The Bax/Bcl-2 ratio in CMA-treated SQUU-B cells was dramatically decreased in comparison with that in the other cell lines treated with CMA. However, when the SQUU-B cells were treated with CMA and a histone deacetylase inhibitor, suberoylanilide hydroxamic acid (SAHA), the SQUU-B cells became more susceptible to the CMA-induced apoptosis. SAHA treatment led to a significantly decrease in the Bcl-2 expression in CMA-treated SQUU-B cells, resulting in a dramatically increased Bax/Bcl-2 ratio in comparison with that observed in the SQUU-B cells treated with CMA alone. These findings suggest that CMA could have an anti-tumor effect on OSCCs. In addition, combination of CMA with other agents, such as SAHA, could help improve the pro-apoptotic effects of CMA even in CMA-resistant OSCC cells.

Oxidative stress induces apoptosis in embryonic cortical neurons.[Pubmed:7903353]

J Neurochem. 1994 Jan;62(1):376-9.

Glutamate-induced glutathione depletion in immature embryonic cortical neurons has been shown to lead to oxidative stress and cell death. We have used this in vitro model to investigate the mechanism(s) by which free radicals induce neuronal degeneration. We find that glutathione depletion leads to hyper-condensation and fragmentation of chromatin into spherical or irregular shapes, a morphologic signature of apoptosis. These morphologic changes are accompanied by laddering of DNA into multiple oligonucleosomal fragments and can be prevented by the antioxidants idebenone and butylated hydroxyanisole. Cell death induced by glutathione depletion can also be prevented by inhibitors of macromolecular synthesis. Taken together, these observations suggest that oxidative stress can induce apoptosis in neurons.

Analysis of single- and double-stranded nucleic acids on polyacrylamide and agarose gels by using glyoxal and acridine orange.[Pubmed:73185]

Proc Natl Acad Sci U S A. 1977 Nov;74(11):4835-8.

We have developed a simple and rapid system for the denaturation of nucleic acids and their subsequent analysis by gel electrophoresis. RNA and DNA are denatured in 1 M glyoxal (ethanedial) and 50% (vol/vol) dimethyl sulfoxide, at 50 degrees. The glyoxalated nucleic acids are then subjected to electrophoresis through either acrylamide or agarose gels in a 10 mM sodium phosphate buffer at pH 7.0. When glyoxalated DNA molecules of known molecular weights are used as standards, accurate molecular weights for RNA are obtained. Furthermore, we have employed the metachromatic stain acridine orange for visualization of nucleic acids in gels. This dye interacts differently with double- and single-stranded polynucleotides, fluorescing green and red, respectively. By using these techniques, native and denatured DNA and RNA molecules can be analyzed on the same slab gel.