SiR-DNA is a far-red, fluorogenic, cell permeable, low background and highly specific probe for DNA. SiR-DNA is based on the DNA minor groove binder bisbenzimide, it allows the labelling of DNA in live cells with high specificity and low background. SiR-DNA is fluorogenic, cell permeable and highly specific for DNA. Sir-DNA stains the nuclei of live cells without the need for genetic manipulation or overexpression. Its emission in the far red minimizes phototoxicity and sample autofluorescence. This reagent has multiple advantages (far-red, fluorogenic, cell permeable, low background and highly specific probe for DNA) over existing probes such Syto61, DRAQ5 or Vybrant DyeCycle Ruby dyes.

SiR-DNA is compatible with GFP and/or m-cherry fluorescent proteins. It can be imaged with standard Cy5 filtersets. SiR-actin can be used for widefield, confocal, SIM or STED imaging in living cells and tissue. Probe quantity allows 50 – 200 staining experiments.*

For comparison information with other nuclear stains click here.

Optical properties

λabs   652 nm

λEm   674 nm
εmax 1.0·105 mol-1·cm-1


*Based on the following conditions: 0.5 – 1 ml staining solution / staining experiments with 0.5 – 1 µM probe concentration. The number of staining experiments can be further increased by reducing volume or probe concentration.

Cytoskeleton, Inc. is the exclusive provider of Spirochrome, Ltd. products in North America.

Live cell time lapse confocal imaging of a HeLa cell stained with SiR-DNA, note the very low background.

For product Datasheets and MSDSs please click on the PDF links below.

For comparison information with other nuclear stains click here.

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Slangen, P L G et al.Protocol for live-cell imaging during Tumor Treating Fields treatment with Inovitro LiveSTAR protocols2022Article Link
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Papini, Diana et al.The Aurora B gradient sustains kinetochore stability in anaphaseCell Reports2021ISSN 2211-1247
Bufe, Anja et al.Wnt signaling recruits KIF2A to the spindle to ensure chromosome congression and alignment during mitosisProceedings of the National Academy of Sciences of the United States of America2021ISSN 1091-6490
Wu, Xi et al.A free-form patterning method enabling endothelialization under dynamic flowBiomaterials2021ISSN 1878-5905
Hilbert, Lennart et al.Transcription organizes euchromatin via microphase separationNature Communications2021ISSN 2041-1723
Ito, Kei K. et al.Cep57 and Cep57L1 maintain centriole engagement in interphase to ensure centriole duplication cycleJournal of Cell Biology2021ISSN 1540-8140
Cordero-Espinoza, Lucía et al.Dynamic cell contacts between periportal mesenchyme and ductal epithelium act as a rheostat for liver cell proliferationCell Stem Cell2021ISSN 1875-9777
Vukušić, Kruno et al.Microtubule-sliding modules based on kinesins EG5 and PRC1-dependent KIF4A drive human spindle elongationDevelopmental Cell2021ISSN 1878-1551
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Warmt, Enrico et al.Differences in cortical contractile properties between healthy epithelial and cancerous mesenchymal breast cellsNew Journal of Physics2021ISSN 1367-2630
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Jagrić, Mihaela et al.Optogenetic control of prc1 reveals its role in chromosome alignment on the spindle by overlap length-dependent forceseLife2021ISSN 2050-084X
Scheffler, Kathleen et al.Two mechanisms drive pronuclear migration in mouse zygotesNature Communications 2021 12:12021ISSN 2041--1723
Buchmann, B. et al.Mechanical plasticity of collagen directs branch elongation in human mammary gland organoidsNature Communications 2021 12:12021ISSN 2041--1723
Geraghty, Zoe et al.The association of Plk1 with the astrin-kinastrin complex promotes formation and maintenance of a metaphase plateJournal of Cell Science2021ISSN 1477-9137
Singh, Divya et al.Destabilization of Long Astral Microtubules via Cdk1-Dependent Removal of GTSE1 from Their Plus Ends Facilitates Prometaphase Spindle OrientationCurrent Biology2021ISSN 1879-0445
Kurzbauer, Marie Therese et al.ATM controls meiotic DNA double-strand break formation and recombination and affects synaptonemal complex organization in plantsThe Plant Cell2021ISSN 1040--4651
Grosser, Steffen et al.Cell and Nucleus Shape as an Indicator of Tissue Fluidity in CarcinomaPhysical Review X2021ISSN 2160-3308
Safaralizade, Mahira et al.Measuring nuclear calcium and actin assembly in living cellsJournal of biochemistry2021ISSN 1756--2651
Zhao, Bing et al.Optogenetic Control of Myocardin-Related Transcription Factor A Subcellular Localization and Transcriptional Activity Steers Membrane Blebbing and Invasive Cancer Cell MotilityAdvanced Biology2021ISSN 2701--0198
Noa, Amra et al.The hierarchical packing of euchromatin domains can be described as multiplicative cascadesPLOS Computational Biology2021ISSN 1553--7358
Menegakis, Apostolos et al.Resistance of Hypoxic Cells to Ionizing Radiation Is Mediated in Part via Hypoxia-Induced QuiescenceCells2021ISSN 2073--4409
Tsuchiya, Kenta et al.Ran-GTP Is Non-essential to Activate NuMA for Mitotic Spindle-Pole Focusing but Dynamically Polarizes HURP Near ChromosomesCurrent Biology2021ISSN 1879-0445
Scott, Stacey J. et al.Evidence that polyploidy in esophageal adenocarcinoma originates from mitotic slippage caused by defective chromosome attachmentsCell Death & Differentiation 2021 28:72021ISSN 1476--5403
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Valenzuela, José Ignacio et al.Localized Intercellular Transfer of Ephrin-As by Trans-endocytosis Enables Long-Term SignalingDevelopmental Cell2020ISSN 1878-1551
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Q1. What are the filter sets for the SiR probes?

A1. The SiR actin and tubulin probes are visualized with standard Cy5 filters.  Optimal excitation is 650 nm and emission is 670 nm.  We recommend filters with an excitation of 630 + 20 nm and an emission of 680 + 20 nm (Fig. 1).  


Figure 1. Excitation (blue) and emission (red) spectra for SiR probes.

Q2. Why do the SiR probes have a low background compared to other fluorophores?

A4. SiR probes are excited by and emit light in the near infrared/far red spectral range, thus avoiding the use of shorter wavelengths such as blue and green light that typically autofluoresce, causing higher background signals.  SiR-coupled probes possess two physical states: 1. a non-fluorescent, closed off-state (spirolactone)  and 2. an open, highly fluorescent on-state (zwitterion).  The binding of the probe to its ligand target favors the highly fluorescent open state while the free unbound probe exists in the closed, non-fluorescent state (Fig. 2).  The fluorescence amplification is 100-fold from the unbound to bound state. This results in a highly sensitive biosensor in which the majority of fluorescence occurs only in the bound state (see Refs. 3 and 4). 


Figure 2. SiR derivatives exist in equilibrium between the fluorescent zwitterionic (open) form (left structure) and the non-fluorescent spiro (closed) form (right structure).

For comparison information with other nuclear stains click here.

Q3: Are the SiR probes stable at room temperature?

A3: Yes, the probes are stable at room temperature for a few days.  However, it strongly depends on the probe and the solvent.  Thus, it is recommended to store all of the probes or solutions at –20°C.


Q4: Are SiR-DNA, SiR-actin and SiR-tubulin toxic to cells?

A4: Yes, above a certain threshold both probes show some effect on cell proliferation and altered actin or microtubule dynamics.  However, the probes are orders of magnitude less toxic than their parent drug.  In HeLa cells, neither actin nor microtubule dynamics were altered at concentrations below 100 nM.  At this concentration, SiR probes efficiently label microtubules and F-actin, allowing for the capture of high signal to noise images.


Q5: Do the probes work on fixed cells?

A5: SiR-DNA labels all types of fixed cells. SiR-actin probes can be used with PFA-fixed cells.  SiR-actin labels F-actin in PFA-fixed cells as efficiently as phalloidin derivatives.  SiR-tubulin labels microtubules only in ethyleneglycol-bis-succinimidyl-succinate (EGS)-fixed cells.  However, a selective labeling of centrosomal microtubules of PFA-fixed cells was observed.  SiR-actin and SiR-tubulin are not suitable for methanol-fixed cells.


Q6: Is it possible to image SiR-probes by STORM?

A6: No—under the very high light intensities typically used in STORM imaging, a phototoxic effect is observed on live cells.


Q7: Which organisms and tissues are stained by SiR-probes?

A7: This list describes only cell lines, tissues, or organisms that have been reported to work.  Omission of a cell line, tissue, or organism does not mean that the SiR-probes will not work with the specific cells, tissues, or organisms.


Homo sapiens: U2OS, fibroblasts, HeLa, HUVEC, MCF-10A, HCT-116, A549, erythrocytes

Mus musculus: C2C12, IA32, skeletal muscle, primary cardiomyocyte, primary oocyte

Rattus norvegicus: primary hippocampal neurons, primary cortical neurons, NRK

Cercopithecus aethiops: COS-7

Mesocricetus auratus: BHK

Drosophila melanogaster: Notum epithelium, S2

Didelphis marsupialis: OK cells


Q8. Do SiR-probes work in 3D cell cultures?

A8: Yes, the probes are able to stain cells in a 3D growth environment.


Q9: What are the correction factors CF260 and CF280 for the SiR fluorophore?

A11: CF260 = 0.116 and CF280  = 0.147

Q10. What is STED microscopy and how does it work?

A10. STED microscopy stands for Stimulated Emission Depletion microscopy.  It is one type of super resolution microscopy which allows the capture of images with a higher resolution than conventional light microscopy which is constrained by diffraction of light.  STED uses 2 laser pulses, one is the excitation pulse which excites the fluorophore, causing it to fluoresce.  The second pulse, referred to as the STED pulse, de-excites the fluorophore via stimulated emission in an area surrounding a central focal spot that is not de-excited and thus continues to fluoresce.  This is accomplished by focusing the STED pulse into a ring shape, a so-called donut, where the center focal spot is devoid of the STED laser pulse, conferring high resolution to the fluorescent area (Fig. 3; see Ref. 1 for more details on STED microscopy).



Figure 3. STED microscopic image of microtubules labeled with SiR-tubulin in human primary dermal fibroblasts.

Q11. Why is the SiR actin (or tubulin) probe good for STED microscopy?

A11. STED microscopy offers the ability to study cellular details on a nanometermolar scale in vivo.  To take advantage of this super resolution microscopy, one must be able to select with high specificity the area to be examined using fluorescent probes.  In addition, the fluorescent probes must be bright, photostable, exhibit no or little phototoxicity, be excited and emit in the far red spectrum.  In addition, if the probe is to be used for live cell imaging (thus avoiding fixation artifacts that occur when cells are fixed), high cell permeability is necessary.  The SiR-DNA, -actin and -tubulin probes fulfill all of these requirements.  In short, the combination of STED and SiR probes allows for unparalleled fluorescent visualization of subcellular actin and tubulin/microtubule structures and their physical characterization in living cells, (see Fig. 4 and Ref. 2). 


Figure 4. STED images of cultured rat hippocampal neurons stained with SiR-actin. Bottom image is a close-up view of part of the top image to clearly visualize actin rings (stripes) with 180 nm periodicity. Courtesy Of Elisa D'Este, MPI Biophysical Chemistry, Göttingen.

For comparison information with other nuclear stains click here.


1. Hell S.W. and Wichmann J. 1994. Breaking the diffraction resolution limit by stimulated emission: stimulated-emission-depletion fluorescence microscopy. Opt. Lett. 19, 780-782.

2. D’Este E. et al. 2015. STED nanoscopy reveals the ubiquity of subcortical cytoskeleton periodicity in living neurons. Cell Rep. 10, 1246-1251.

3. Lukinavicius G. et al. 2013. A near-infrared fluorophore for live-cell super-resolution microscopy of cellular proteins. Nat. Chem. 5, 132-139.

4. Lukinavicius G. et al. 2014. Fluorogenic probes for live-cell imaging of the cytoskeleton. Nature Methods. 11, 731-733.