Acti-stain 555 phalloidin

$0.00

SKU

PHDH1

Datasheet

MSDS

Amount

300 Slides (Cat. # PHDH1-A) + $232.00

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Details

Product Uses Include

Fluorescent staining of actin filaments in fixed tissue sections and tissue culture cells preparations.Note: Unlike many actin antibodies, Acti-stain™ 555 phalloidin binds only to F-actin resulting in low background fluorescence. Furthermore, actin staining is not appreciably different between species.
Preparation of stabilized fluorescent actin filaments in vitro.

Actin staining is very useful in determining the structure and function of the cytoskeleton in living a nd fixed cells. The actin cytoskeleton is a very dynamic and labile structure in the living cell, but it can be fixed with paraformaldehyde prior to probing or staining for actin structures.

Material

Phalloidin is a seven amino acid peptide toxin from the mushroom Amanita phalloides, which binds specifically and with high affinity (Kd 20 nM) to the polymerized form of actin (F-actin). Phalloidin lowers the critical concentration of actin polymerization to less than 1 µg/ml, thereby acting as a polymerization enhancer. Phalloidin has been labeled with a proprietary red fluorescent dye which allows it to be used to stain actin filaments in tissue cultured cells and tissue sections (1, see Figure 1) and cell-free preparations. Acti-stain™ 555 phalloidin-labeled actin filaments retain many functional characteristics of unlabeled actin including their ability to interact with myosin. Actin-stain™ 555 phalloidin is supplied as a red solid.

Note: Phalloidin is toxic and must be handled with care (LD50 human = 2mg/Kg).

Click Here for Detailed Methods

Example Results and Specifications

Figure 1. Actin Stress Fibers stained with Acti-stain™ 555 in a Swiss 3T3 cell.

Swiss 3T3 cell with activated Rho, nucleus is stained with Dapi (blue) and F-actin is stained with Acti-stain™ 555 (red F-actin, Cat.# PHDH1).

Figure 2. Emission and excitation scans for Acti-stain™ phalloidins

Absorbance and fluorescence scan of Acti-stain™ 555. Labeled phalloidin was diluted into methanol and its absorbance and excitation spectra were scanned between 300-750 and 560-750 nm, respectively. Absorbance peaks at 560 nm and fluorescence at 575 nm.

Acti-stain phalloidins are the most well characterized phalloidins available. Tabe 1 describes their brightness, photostability, background and affinity constants to F-actin. Compare these performance characteristics to other fluorescent phalloidins and you will see the advantages of using Acti-stain™ for your actin staining requirements.

Table 1. Biochemical characteristics of fluorescent phalloidins

Conjugate	Cat.#	Wavelengths (Ex/Em)	*Brightness (AFU)**	Stability to photobleaching(half life in seconds)**	Background (% of total AFU at 100nM)**	Affinity (Kd in nanomolar)
Fluorescein-phalloidin	na	485/535 FITC filter set	432	6	22	72 +/-12
Acti-stain™ 488	PHDG1	485/535 FITC filter set	832	57	5	55 +/-8
Acti-stain™ 535	PHDR1	535/585 TRITC filter set	430	27	12	36 +/-7
Acti-stain™ 555	PHDH1	535/585 TRITC filter set	551	46	16	63 +/-8
Acti-stain™ 670	PHDN1	640/680 Cy5 filter set	332	8	18	50 +/-12

* = AFU's measured by quantitative cell imaging. ** = Measured in stained Swiss 3T3 cells in the absence of antifade.

For more information about protocols to fix and stain cells click here.

References

1. Wulf, E. et al. (1979). Proc Natl Acad Sci USA. 76(9):4498-4502.

2. Kron, S.J. et al. (1991). Meth. Enzmol. 196: 399-416.

About

For product Datasheets and MSDSs please click on the PDF links below. For additional information, click on the FAQs tab above or contact our Technical Support department at tservice@cytoskeleton.com

Citations

Author	Title	Journal	Year	Article Link
Huang, Qichao et al.	TFAM loss induces nuclear actin assembly upon mDia2 malonylation to promote liver cancer metastasis	The EMBO Journal	2022	ISSN 0261--4189
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Schimmel, Lilian et al.	Endothelial cells are not productively infected by SARS-CoV-2	Clinical and Translational Immunology	2021	ISSN 2050-0068
Kimura, Ken ichi et al.	Crucial role of framework with cytoskeletal actin filaments for shaping microstructure of footpad setae in the ladybird beetle, Harmonia axyridis	Arthropod Structure and Development	2021	ISSN 1873-5495
Schiweck, Juliane et al.	Drebrin controls scar formation and astrocyte reactivity upon traumatic brain injury by regulating membrane trafficking	Nature Communications	2021	ISSN 2041-1723
Gharanei, Seley et al.	Vascular Adhesion Protein-1 Determines the Cellular Properties of Endometrial Pericytes	Frontiers in Cell and Developmental Biology	2021	ISSN 2296-634X
Goering, Jeremy P. et al.	In-frame deletion of SPECC1L microtubule association domain results in gain-of-function phenotypes affecting embryonic tissue movement and fusion events	Human molecular genetics	2021	ISSN 1460-2083
Shindo, Taiki et al.	Visualization of endogenous nuclear F-actin in mouse embryos reveals abnormal actin assembly after somatic cell nuclear transfer	Journal of biochemistry	2021	ISSN 1756-2651
Wurzer, Hannah et al.	Intrinsic Resistance of Chronic Lymphocytic Leukemia Cells to NK Cell-Mediated Lysis Can Be Overcome In Vitro by Pharmacological Inhibition of Cdc42-Induced Actin Cytoskeleton Remodeling	Frontiers in Immunology	2021	ISSN 1664-3224
Kanamoto, Takashi et al.	Integrin α2β1 plays an important role in the interaction between human articular cartilage-derived chondrocytes and atelocollagen gel	Scientific Reports	2021	ISSN 2045-2322
Liang, Fei et al.	Sorbs2 is a genetic factor contributing to cardiac malformation of 4q deletion syndrome patients	eLife	2021	ISSN 2050-084X
Haines, Adam et al.	Cross Talk between ARF1 and RhoA Coordinates the Formation of Cytoskeletal Scaffolds during Chlamydia Infection	mBio	2021	ISSN 2150-7511
Liang, Wei et al.	Necroptosis activates UPR sensors without disrupting their binding with GRP78	Proceedings of the National Academy of Sciences of the United States of America	2021	ISSN 1091-6490
Nanamatsu, Azuma et al.	Vasopressin Induces Urinary Uromodulin Secretion by Activating PKA (Protein Kinase A)	Hypertension	2021	ISSN 1524-4563
Woo, Marcel S. et al.	Neuronal metabotropic glutamate receptor 8 protects against neurodegeneration in CNS inflammation	Journal of Experimental Medicine	2021	ISSN 1540-9538
Kimura, Ken ichi et al.	Framework with cytoskeletal actin filaments forming insect footpad hairs inspires biomimetic adhesive device design	Communications Biology	2020	ISSN 2399-3642
Sancey, Lucie et al.	Multiparametric investigation of non functionalized-AGuIX nanoparticles in 3D human airway epithelium models demonstrates preferential targeting of tumor cells	Journal of Nanobiotechnology	2020	ISSN 1477-3155
Campisi, Marco et al.	Tumor-Derived cGAMP Regulates Activation of the Vasculature	Frontiers in Immunology	2020	ISSN 1664-3224
Chemla, Yoav et al.	Carbon nanostructures as a scaffold for human embryonic stem cell differentiation toward photoreceptor precursors	Nanoscale	2020	ISSN 2040--3372
Bokelmann, Marcel et al.	Utility of primary cells to examine npc1 receptor expression in mops condylurus, a potential ebola virus reservoir	PLoS Neglected Tropical Diseases	2020	ISSN 1935-2735
Fu, Yi et al.	Elevation of JAML Promotes Diabetic Kidney Disease by Modulating Podocyte Lipid Metabolism	Cell Metabolism	2020	ISSN 1932-7420
Ding, Dalian et al.	Antioxidative stress-induced damage in cochlear explants	Journal of Otology	2020	ISSN 1672-2930
Chen, Lena et al.	Ankyrin-B p.S646F undergoes increased proteasome degradation and reduces cell viability in the H9c2 rat ventricular cardiomyoblast cell line		2020	PMID 31965814
Shi, Rui et al.	Rapid Osseointegration Bestowed by Carbonate Apatite Coating of Rough Titanium	Advanced Materials Interfaces	2020	ISSN 2196-7350
Ding, Dalian et al.	Some Ototoxic Drugs Destroy Cochlear Support Cells Before Damaging Sensory Hair Cells	Neurotoxicity Research	2020	ISSN 1476-3524
Chen, Lixia et al.	CSRP2 suppresses colorectal cancer progression via p130Cas/Rac1 axis-meditated ERK, PAK, and HIPPO signaling pathways	Theranostics	2020	ISSN 1838-7640
Liu, Yi et al.	Effect of F-actin and Microtubules on Cellular Mechanical Behavior Studied Using Atomic Force Microscope and an Image Recognition-Based Cytoskeleton Quantification Approach	International Journal of Molecular Sciences	2020	ISSN 1422-0067
Pęziński, Marcin et al.	An improved method for culturing myotubes on laminins for the robust clustering of postsynaptic machinery	Scientific Reports	2020	ISSN 2045-2322
Hamdan, Hamdan et al.	Mapping axon initial segment structure and function by multiplexed proximity biotinylation	Nature Communications	2020	ISSN 2041-1723
Hall, Everett G. et al.	SPECC1L regulates palate development downstream of IRF6	Human Molecular Genetics	2020	ISSN 1460-2083
Vieira, Tânia et al.	Electrospun biodegradable chitosan based-poly(urethane urea) scaffolds for soft tissue engineering	Materials Science and Engineering C	2019	ISSN 1873-0191
Fultz, Robert et al.	Phagocytosis by macrophages depends on histamine H2 receptor signaling and scavenger receptor 1	MicrobiologyOpen	2019	ISSN 2045-8827
Vohnoutka, Rishel B. et al.	The focal adhesion scaffold protein Hic-5 regulates vimentin organization in fibroblasts	Molecular Biology of the Cell	2019	ISSN 1939-4586
Ye, Haibo et al.	Bilirubin-induced neurotoxic and ototoxic effects in rat cochlear and vestibular organotypic cultures	NeuroToxicology	2019	ISSN 1872-9711
Siddiqui, Nida et al.	PTPN21 and Hook3 relieve KIF1C autoinhibition and activate intracellular transport	Nature Communications	2019	ISSN 2041-1723
Ramos, Ana Raquel et al.	Lipid phosphatases SKIP and SHIP2 regulate fibronectin-dependent cell migration in glioblastoma	FEBS Journal	2019	ISSN 1742-4658
Ueta, Miho et al.	Effects of TGF-β1 on the migration and morphology of RAW264.7 cells in vitro	Molecular Medicine Reports	2019	ISSN 1791-3004
Mandigo, Torrey R. et al.	Drosophila emerins control LINC complex localization and transcription to regulate myonuclear position	Journal of Cell Science	2019	ISSN 1477-9137
Nishiyama, Keita et al.	A new approach for analyzing an adhesive bacterial protein in the mouse gastrointestinal tract using optical tissue clearing	Scientific Reports	2019	ISSN 2045-2322
Ferguson, Sylvia H. et al.	Interferon-λ3 Promotes Epithelial Defense and Barrier Function Against Cryptosporidium parvum Infection	Cmgh	2019	ISSN 2352-345X
Brauer, Philip R. et al.	Krüppel-like factor 4 mediates cellular migration and invasion by altering RhoA activity	Cell Communication and Adhesion	2018	ISSN 1543-5180
Ghosh, Somadri et al.	Inhibition of SHIP2 activity inhibits cell migration and could prevent metastasis in breast cancer cells	Journal of Cell Science	2018	ISSN 1477-9137
Camuglia, Jaclyn M. et al.	An RNAi based screen in Drosophila larvae identifies fascin as a regulator of myoblast fusion and myotendinous junction structure	Skeletal Muscle	2018	ISSN 2044-5040
Bardai, Farah H. et al.	Lrrk promotes tau neurotoxicity through dysregulation of actin and mitochondrial dynamics	PLoS Biology	2018	ISSN 1545-7885
Wang, Liqun et al.	Tissue and cellular rigidity and mechanosensitive signaling activation in Alexander disease	Nature Communications	2018	ISSN 2041-1723
Bardai, Farah H. et al.	A conserved cytoskeletal signaling cascade mediates neurotoxicity of FTDP-17 tau mutations in vivo	Journal of Neuroscience	2018	ISSN 1529-2401
Del Signore, Steven J. et al.	The WAVE Regulatory Complex and Branched F-Actin Counterbalance Contractile Force to Control Cell Shape and Packing in the Drosophila Eye	Developmental Cell	2018	ISSN 1878-1551
Della Torre, Camilla et al.	Adsorption of B(α)P on carbon nanopowder affects accumulation and toxicity in zebrafish (Danio rerio) embryos	Environmental Science: Nano	2017	ISSN 2051-8161
Nakamura, Koji et al.	Exosomes promote ovarian cancer cell invasion through transfer of CD44 to peritoneal mesothelial cells	Molecular Cancer Research	2017	ISSN 1557-3125
Girolimetti, Giulia et al.	Platinum-induced mitochondrial DNA mutations confer lower sensitivity to paclitaxel by impairing tubulin cytoskeletal organization	Human Molecular Genetics	2017	ISSN 1460-2083
Itano, Seiji et al.	Colchicine attenuates renal fibrosis in a murine unilateral ureteral obstruction model	Molecular Medicine Reports	2017	ISSN 1791-3004
Collins, Mary Ann et al.	Emery-Dreifuss muscular dystrophy-linked genes and centronuclear myopathy-linked genes regulate myonuclear movement by distinct mechanisms	Molecular Biology of the Cell	2017	ISSN 1939-4586
Ando, Fumiaki et al.	Wnt5a induces renal AQP2 expression by activating calcineurin signalling pathway	Nature Communications	2016	ISSN 2041-1723
Dorland, Yvonne L. et al.	The F-BAR protein pacsin2 inhibits asymmetric VE-cadherin internalization from tensile adherens junctions	Nature Communications	2016	ISSN 2041-1723
Zeng, Fa Min et al.	Biological characterization of three immortalized esophageal epithelial cell lines	Molecular Medicine Reports	2016	ISSN 1791-3004
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Burks, J. et al.	ISGylation governs the oncogenic function of Ki-Ras in breast cancer	Oncogene 2014 33:6	2013	ISSN 1476--5594
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Faqs

Question 1: Can I use fluorescently-labeled phalloidin to stain F-actin in living cells?

Answer 1: Unfortunately, no, phalloidin cannot be used to stain F-actin in living cells. Phalloidins are used to stain F-actin in fixed cells. Fluorescent phalloidins only bind to the native quaternary structure of F-actin which provides a low background. The correct fixation condition for phalloidin binding is 3.7% (v/v) paraformaldehyde in PBS for 10 min because it retains the quaternary protein structure which is necessary for high affinity binding of phalloidin. Methanol fixation destroys the native conformation and hence is not suitable for F-actin staining with phalloidin. To monitor actin dynamics in living cells, micro-injection of rhodamine-labeled actin (Cat. # APHR or AR05) is recommended. Please see those datasheets for more information.

Question 2: Which of the fluorescently-labeled phalloidin is the most stable/brightest?

Answer 2: The brightest and most stable of the Acti-stains is Acti-stain 488 (green fluorescence; Cat. # PHDG1). Please see the table below for additional information on all of our Acti-stains.

Conjugate	Cat. #	Wavelengths (Ex/EM)	Brightness (AFU)	*Stability to photobleaching (1/2 life in sec)**	Background (% of total AFU at 100 nM)
Acti-stain™ 488	PHDG1	485/535	832	57	5
Acti-stain™ 535	PHDR1	535/585	430	27	12
Acti-stain™ 555	PHDH1	535/585	551	46	16
Acti-stain™ 670	PHDN1	640/680	332	18	18

* = measured in the absence of antifade.

If you have any questions concerning this product, please contact our Technical Service department at tservice@cytoskeleton.com.

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Acti-stain 555 phalloidin

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