Unique spectral signatures of the nucleic acid dye acridine orange can distinguish cell death by apoptosis and necroptosis

Cellular injury and death are ubiquitous features of disease, yet tools to detect them are limited and insensitive to subtle pathological changes. Acridine orange (AO), a nucleic acid dye with unique spectral properties, enables real-time measurement of RNA and DNA as proxies for cell viability during exposure to various noxious stimuli. This tool illuminates spectral signatures unique to various modes of cell death, such as cells undergoing apoptosis versus necrosis/necroptosis. This new approach also shows that cellular RNA decreases during necrotic, necroptotic, and apoptotic cell death caused by demyelinating, ischemic, and traumatic injuries, implying its involvement in a wide spectrum of tissue pathologies. Furthermore, cells with pathologically low levels of cytoplasmic RNA are detected earlier and in higher numbers than with standard markers including TdT-mediated dUTP biotin nick-end labeling and cleaved caspase 3 immunofluorescence. Our technique highlights AO-labeled cytoplasmic RNA as an important early marker of cellular injury and a sensitive indicator of various modes of cell death in a range of experimental models.

• Publication year

  2017

• Venue

  Journal of Cell Biology

• Publication date

  2017-04-03

• Fields of study

  Biology, Medicine

• Identifiers
  DOI 10.1083/jcb.201602028PMID 28264914PMCID 5379938
• External record

  Open on Semantic Scholar

• Source metadata

  Semantic Scholar, PubMed

• No claims are published for this paper.

• No concepts are published for this paper.

• CHIP controls necroptosis through ubiquitylation- and lysosome-dependent degradation of RIPK3

  2016cited by this paper
• Experimental Demyelination and Remyelination of Murine Spinal Cord by Focal Injection of Lysolecithin

  2015cited by this paper
• Activation of necroptosis in multiple sclerosis.

  2015cited by this paper
• An RNA-Sequencing Transcriptome and Splicing Database of Glia, Neurons, and Vascular Cells of the Cerebral Cortex

  2014cited by this paper
• Necroptosis.

  2014cited by this paper
• Remyelination after spinal cord injury: is it a target for repair?

  2014cited by this paper
• Determination of apoptotic and necrotic cell death in vitro and in vivo.

  2013cited by this paper
• TNF can activate RIPK3 and cause programmed necrosis in the absence of RIPK1

  2013cited by this paper
• The proteases HtrA2/Omi and UCH-L1 regulate TNF-induced necroptosis

  2013cited by this paper
• Regulation of cytoplasmic mRNA decay

  2012cited by this paper
• Apoptosis-like programmed cell death induces antisense ribosomal RNA (rRNA) fragmentation and rRNA degradation in Leishmania

  2012influential reference
• Necroptosis

  2012cited by this paper
• Dectin-1 is an extracellular pathogen sensor for the induction and processing of IL-1β via a noncanonical caspase-8 inflammasome

  2012cited by this paper
• Nuclear bodies: multifunctional companions of the genome.

  2012cited by this paper
• Regulation of cytoplasmic mRNA decay

  2012cited by this paper
• Biogenesis and function of nuclear bodies.

  2011cited by this paper
• Derivation of Enriched Oligodendrocyte Cultures and Oligodendrocyte/Neuron Myelinating Co-cultures from Post-natal Murine Tissues

  2011cited by this paper
• CXCR2-positive neutrophils are essential for cuprizone-induced demyelination: relevance to multiple sclerosis

  2010cited by this paper
• Multiple sclerosis: Distribution of inflammatory cells in newly forming lesions

  2009cited by this paper
• The cuprizone animal model: new insights into an old story

  2009cited by this paper
• Apoptosis and necrosis: detection, discrimination and phagocytosis.

  2008cited by this paper
• Real-time measurement of free Ca2+ changes in CNS myelin by two-photon microscopy

  2007cited by this paper
• Morphological and cytochemical determination of cell death by apoptosis

  2007cited by this paper
• Towards standardization of RNA quality assessment using user-independent classifiers of microcapillary electrophoresis traces

  2005cited by this paper
• Chemical inhibitor of nonapoptotic cell death with therapeutic potential for ischemic brain injury

  2005cited by this paper
• Energy constraints on the evolution of gene expression.

  2005cited by this paper
• Luminescence of the solid complexes of acridine orange with RNA.

  2005cited by this paper
• Developmental differences in H2O2‐induced oligodendrocyte cell death: role of glutathione, mitogen‐activated protein kinases and caspase 3

  2004cited by this paper
• Increased expression of Nkx2.2 and Olig2 identifies reactive oligodendrocyte progenitor cells responding to demyelination in the adult CNS.

  2004cited by this paper
• Developmental differences in HO-induced oligodendrocyte cell death: role of glutathione, mitogen-activated protein kinases and caspase 3.

  2004cited by this paper
• Relapsing and remitting multiple sclerosis: Pathology of the newly forming lesion

  2004cited by this paper
• Translation inhibition during the induction of apoptosis: RNA or protein degradation?

  2004influential reference
• Reasons to reconsider the significance of apoptosis for cancer therapy

  2003cited by this paper
• Mitochondrial Complex I Inhibitor Rotenone Induces Apoptosis through Enhancing Mitochondrial Reactive Oxygen Species Production*

  2003cited by this paper
• Degradation of cellular mRNA is a general early apoptosis‐induced event

  2002cited by this paper
• Improved Detection of Apoptotic Cells in Archival Paraffin Sections: Immunohistochemistry Using Antibodies to Cleaved Caspase 3

  2002cited by this paper
• MIP-1α, MCP-1, GM-CSF, and TNF-α Control the Immune Cell Response That Mediates Rapid Phagocytosis of Myelin from the Adult Mouse Spinal Cord

  2001cited by this paper
• MIP-1alpha, MCP-1, GM-CSF, and TNF-alpha control the immune cell response that mediates rapid phagocytosis of myelin from the adult mouse spinal cord.

  2001cited by this paper
• Oligodendroglial apoptosis occurs along degenerating axons and is associated with FAS and p75 expression following spinal cord injury in the rat.

  2001cited by this paper
• Identification of Apoptotic Cells by Formamide-induced DNA Denaturation in Condensed Chromatin

  2001cited by this paper
• Temporal-spatial pattern of acute neuronal and glial loss after spinal cord contusion.

  2001cited by this paper
• The Neurotoxicant, Cuprizone, as a Model to Study Demyelination and Remyelination in the Central Nervous System

  2001cited by this paper
• Insulin-Like Growth Factor-1 Inhibits Mature Oligodendrocyte Apoptosis during Primary Demyelination

  2000cited by this paper
• Lysophosphatidylcholine induces rapid recruitment and activation of macrophages in the adult mouse spinal cord

  2000cited by this paper
• Mature oligodendrocyte apoptosis precedes IGF‐1 production and oligodendrocyte progenitor accumulation and differentiation during demyelination/remyelination

  2000cited by this paper
• 28S ribosome degradation in lymphoid cell apoptosis: evidence for caspase and Bcl-2-dependent and -independent pathways

  2000cited by this paper
• Distinctive patterns of PDGF-A, FGF-2, IGF-I, and TGF-beta1 gene expression during remyelination of experimentally-induced spinal cord demyelination.

  1999cited by this paper
• Titrating the Effects of Mitochondrial Complex I Impairment in the Cell Physiology*

  1999cited by this paper
• Differential responses of oligodendrocytes to tumor necrosis factor and other pro‐apoptotic agents: Role of ceramide in apoptosis

  1999cited by this paper
• Apoptotic Cells Associated with Wallerian Degeneration After Experimental Spinal Cord Injury: A Possible Mechanism of Oligodendroglial Death

  1999cited by this paper
• Apoptosis after traumatic human spinal cord injury.

  1998cited by this paper
• Inhibition of Caspases Increases the Sensitivity of L929 Cells to Necrosis Mediated by Tumor Necrosis Factor

  1998cited by this paper
• Intracellular Calcium and Cell Death during Ischemia in Neonatal Rat White Matter Astrocytes In Situ

  1998cited by this paper
• Mode of cell injury and death after hydrogen peroxide exposure in cultured oligodendroglia cells.

  1998cited by this paper
• Apoptosis and delayed degeneration after spinal cord injury in rats and monkeys

  1997influential reference
• Neuronal and Glial Apoptosis after Traumatic Spinal Cord Injury

  1997influential reference
• Accumulation of Sequence-specific RNA-binding Proteins in the Cytosol of Activated T Cells Undergoing RNA Degradation and Apoptosis (*)

  1995influential reference
• MORPHOLOGICAL AND CYTOCHEMICAL STUDIES OF THE

  1992cited by this paper
• Identification of programmed cell death in situ via specific labeling of nuclear DNA fragmentation

  1992cited by this paper
• Glucocorticoids stimulate ribonucleic acid degradation in isolated rat thymic lymphocytes in vitro.

  1982cited by this paper
• The fluorescence of dye-nucleic acid complexes☆

  1981cited by this paper
• Cell death: the significance of apoptosis.

  1980cited by this paper
• Central nervous system demyelination and remyelination in the mouse: an ultrastructural study of cuprizone toxicity.

  1978cited by this paper
• Simultaneous staining of ribonucleic and deoxyribonucleic acids in unfixed cells using acridine orange in a flow cytofluorometric system.

  1977cited by this paper
• The effect of injections of lysophosphatidyl choline into white matter of the adult mouse spinal cord.

  1972cited by this paper
• Fluorescence-excitation spectra of acridine orange-DNA and -RNA systems

  1967cited by this paper

• Schisandrol A induces cardiac developmental toxicity in zebrafish (Danio rerio) via disruption of calcium homeostasis, oxidative stress, and apoptosis.

  2026cites this paper
• Vision of Life's Code: Molecular Probe Nanoarchitectonics for Deep RNA/DNA Illumination.

  2026cites this paper
• Targeting NgR1/NogoA Signalling Enhances Natural Killer Cell Cytotoxicity and Alters Immune and Oncogenic Pathways in TNBC Cells

  2026cites this paper
• Acousto-holographic investigation of the changes in morphology and cellular biomechanics under oxidative stress

  2026cites this paper
• Viability-Informed Single-Cell Mass Spectrometry for More Comprehensive Metabolic Analysis.

  2026cites this paper
• Pristimerin Alleviates Nanoparticulate Titanium Dioxide-Triggered Neurodegeneration in Hippocampal and Cortex by Downregulation of Inflammation and Oxidative Damage.

  2026cites this paper
• Protective Role and Mechanism of Flavonoids from Smilax glabra Roxb. against Lead-Induced Neurotoxicity and Hematotoxicity in Danio rerio.

  2026cites this paper
• Viable and Functional: Long-Term −80 °C Cryopreservation Sustains CD34+ Integrity and Transplant Success

  2025cites this paper
• Non-canonical role of ''S6K1–SGK1'' pathway in neuronal necroptosis following traumatic brain injury

  2025cites this paper
• Ecotoxicity of microplastics in embryos and newly hatched snails of Biomphalaria glabrata

  2025cites this paper
• Encapsulation of Curcumin and Gemcitabine: Cytotoxic Effect and Mechanisms of Death in Lung Cancer

  2025cites this paper
• An optimized method for estimating toxicity and cell death in the mysid model using acridine orange staining

  2025cites this paper
• Serum-Stable, Cationic, α-Helical AMPs to Combat Infections of ESKAPE Pathogens and C. albicans.

  2025cites this paper
• ACC-1 as a Possible Biochemical Indicator of Lipoapoptosis in In Vivo and In Vitro Models of MAFLD

  2025cites this paper
• Rapid slide-free histopathology via fluorescent layer-adaptive subcellular imaging: clinical-accessible platform for diverse tissues

  2025cites this paper
• Spectral Phasor Analysis for Hyperspectral Imaging to elucidate Acridine Orange Photophysics in Cells

  2025influential citation
• Evolution of Physicochemical and Toxicological Properties During Aqueous Exposition of the Nanoparticulated Food Colours E-174 and E-172: Study in Caenorhabditis elegans

  2025cites this paper
• Design and optimization of thermosensitive injectable alginate-based hydrogels: potential for loading therapeutic compounds

  2025cites this paper
• Biosynthesized bimetallic nanoparticles containing CeO2 and ZnO exert shape and size dependent anticancer effects

  2025cites this paper
• Age-impaired remyelination is associated with dysregulated microglial transitions

  2025cites this paper
• Visualizing the chronicle of multiple cell fates using a near-IR dual-RNA/DNA–targeting probe

  2025cites this paper
• Acellular Human Amniotic Membrane: A Safe and Stable Matrix for Corneal Stromal Regeneration

  2024cites this paper
• Detection of dsRNA by Acridine Orange Staining.

  2024cites this paper
• A monofunctional Pt(II) complex combats triple negative breast cancer by triggering lysosome-dependent cell death.

  2024cites this paper
• Dimethyl fumarate induces cardiac developmental toxicity in zebrafish via down-regulation of oxidative stress.

  2024cites this paper
• Qingfei Yin alleviates Streptococcus pneumoniae pneumonia by promoting complete autophagy to suppress necroptosis.

  2024cites this paper
• Enhancing the porosity of biphasic calcium phosphate using polyethylene glycol as the porogen for bone regeneration applications

  2024cites this paper
• Nucleic acid-binding bis-acridine orange dyes with improved properties for bioimaging and PCR applications.

  2024cites this paper
• Protocol for detecting lysosome quantity and membrane permeability in acute myeloid leukemia cell lines

  2024cites this paper
• Cellular Output and Physicochemical Properties of the Membrane-Derived Vesicles Depend on Chemical Stimulants

  2024cites this paper
• Fly Ash Concrete Specimens Admixed with Nanoparticles and Their Interaction with Seawater

  2024cites this paper
• Toxic Features and Metabolomic Intervention of Glabrene, an Impurity Found in the Pharmaceutical Product of Glabridin

  2024cites this paper
• Acridine Orange and Its Removal from Aqueous Solutions by Adsorption

  2024cites this paper
• Fluorescent probes for neuroscience: imaging ex vivo brain tissue sections.

  2024cites this paper
• Bacteriophage therapy reduces Staphylococcus aureus in a porcine and human ex vivo burn wound infection model

  2024influential citation
• Naringenin Mitigates Nanoparticulate-Aluminium Induced Neuronal Degeneration in Brain Cortex and Hippocampus through Downregulation of Oxidative Stress and Neuroinflammation.

  2024cites this paper
• Microglial phagocytosis of single dying oligodendrocytes is mediated by CX3CR1 but not MERTK.

  2024cites this paper
• Avoiding cantharidin through ionotropic receptors.

  2024cites this paper
• Study of the Early Effects of Chitosan Nanoparticles with Glutathione in Rats with Osteoarthrosis

  2023cites this paper
• miR-19-3p Targets PTEN to Regulate Cervical Cancer Cell Proliferation, Invasion, and Autophagy

  2023cites this paper
• Low-level laser selectively inhibiting colorectal cancer cell metabolic activity and inducing apoptosis for delaying the development of intestinal cancer

  2023cites this paper
• Isolation and structural characterization of exopolysaccharide from the Cordyceps cicadae and the immunomodulatory activity on RAW264.7 cells

  2023cites this paper
• Comparative analysis of α-pinene alone and combined with temozolomide in human glioblastoma cells

  2023cites this paper
• Oligodendrocyte Maturation Alters the Cell Death Mechanisms That Cause Demyelination

  2023cites this paper
• Polystyrene nanoplastics induce developmental impairments and vasotoxicity in zebrafish (Danio rerio).

  2023cites this paper
• Microglial phagocytosis of single dying oligodendrocytes is mediated by CX3CR1 but not MERTK

  2023cites this paper
• Bone marrow mesenchymal stem cells and exercise restore motor function following spinal cord injury by activating PI3K/AKT/mTOR pathway

  2022cites this paper
• A novel pH- and glutathione-responsive drug delivery system based on in situ growth of MOF199 on mesoporous organic silica nanoparticles targeting the hepatocellular carcinoma niche

  2022cites this paper
• Effect of Coated Silver Nanoparticles on Cancerous vs. Healthy Cells

  2022cites this paper
• Methodological advances in necroptosis research: From challenges to solutions

  2022cites this paper
• Oligodendrocyte death and myelin loss in the cuprizone model: an updated overview of the intrinsic and extrinsic causes of cuprizone demyelination

  2022cites this paper
• Nerolidol induced apoptosis via PI3K/JNK regulation through cell cycle arrest in MG‐63 osteosarcoma cells

  2022cites this paper
• Bi-Functional Gold Nanorod-Protein Conjugates with Biomimetic BSA@Folic Acid Corona for Improved Tumor Targeting and Intracellular Delivery of Therapeutic Proteins in Colon Cancer 3D Spheroids.

  2022cites this paper
• Acridine-based fluorophores with improved lightfastness properties

  2022cites this paper
• Single-cell microglial transcriptomics during demyelination defines a microglial state required for lytic carcass clearance

  2022influential citation
• In-Vitro Antibacterial and Anti-Inflammatory Effects of Surfactin-Loaded Nanoparticles for Periodontitis Treatment

  2021cites this paper
• Content of Nucleic Acids in the Adhesive T-Lymphoblast Jurkat Line and Their Mobility In Vitro

  2021cites this paper
• TNF-α-dependent neuronal necroptosis regulated in Alzheimer's disease by coordination of RIPK1-p62 complex with autophagic UVRAG

  2021influential citation
• PNKP is required for maintaining the integrity of progenitor cell populations in adult mice

  2021cites this paper
• Alcohol‐induced aggression in Drosophila

  2021cites this paper
• Curcumin Protects HepG2 Cells from the Cytotoxic Effect of Drugs with Anti-fibrotic Activity

  2021cites this paper
• Genotoxicity of chloroacetamide herbicides and their metabolites in vitro and in vivo

  2021cites this paper
• An Automated Differential Nuclear Staining Assay for Accurate Determination of Mitocan Cytotoxicity.

  2020cites this paper
• Functional State of Mesenchymal Stem Cells upon Exposure to Bioactive Coatings on Titanium Alloys

  2020cites this paper
• Eliciting OTUD3/RIPK-Dependent Necroptosis to Prevent Epithelial Ovarian Cancer

  2020cites this paper
• Acridine Orange: A Review of Novel Applications for Surgical Cancer Imaging and Therapy

  2019cites this paper
• Dendritic Cell-Targeted pH-Responsive Extracellular Vesicles for Anticancer Vaccination

  2019cites this paper
• Intersections between Regulated Cell Death and Autophagy.

  2019cites this paper
• Diltiazem potentiation of doxorubicin cytotoxicity and cellular uptake in human breast cancer cells

  2019cites this paper
• Measurement and models accounting for cell death capture hidden variation in compound response

  2019cites this paper
• Simple Histochemical Methods to Detect Cell Death in the Eye-Antennae Imaginal Disc of Drosophila

  2019cites this paper
• Evaluation of Fluorescent Stains for Viability Assessment of the Potato Cyst Nematodes Globodera pallida and G. ellingtonae

  2019cites this paper
• Effects of ethanol intake on anti-oxidant responses and the lifespan of Caenorhabditis elegans

  2019cites this paper
• Mechanisms of lysophosphatidylcholine‐induced demyelination: A primary lipid disrupting myelinopathy

  2018cites this paper
• cology combined with functional metabolomics discover bile acid metabolism as a promising target for mirabilite against colorectal cancer †

  2018cites this paper
• INFLUENCE OF BIOCOMPATIBLE COATINGS ON MAGNESIUM ON THE FUNCTIONAL STATE OF MESENCHYMAL STEM CELLS

  2018cites this paper
• Network pharmacology combined with functional metabolomics discover bile acid metabolism as a promising target for mirabilite against colorectal cancer

  2018cites this paper
• Matrix-Assisted Regulation of Antimicrobial Properties: Mechanistic Elucidation with Ciprofloxacin-Based Polymeric Hydrogel Against Vibrio Species.

  2018cites this paper
• Wash-free instant detection of giant plasma membrane vesicles.

  2018cites this paper
• Inorganic fullerene-like molybdenum selenide with good biocompatibility synthesized by laser ablation in liquids

  2018cites this paper
• Visualization of murine lymph vessels using photoacoustic imaging with contrast agents

  2018cites this paper
• Ablative Hypofractionated Radiation Therapy Enhances Non-Small Cell Lung Cancer Cell Killing via Preferential Stimulation of Necroptosis In Vitro and In Vivo.

  2018cites this paper
• Conserved nucleolar stress at the onset of cell death

  2017cites this paper