Phalloidin-iFluor® 647 Conjugate
Ordering information
Price | |
Catalog Number | |
Unit Size | |
Quantity |
Additional ordering information
Telephone | 1-800-990-8053 |
Fax | 1-800-609-2943 |
sales@aatbio.com | |
International | See distributors |
Bulk request | Inquire |
Custom size | Inquire |
Shipping | Standard overnight for United States, inquire for international |
Physical properties
Molecular weight | 1408.65 |
Solvent | DMSO |
Spectral properties
Correction Factor (260 nm) | 0.03 |
Correction Factor (280 nm) | 0.03 |
Correction Factor (656 nm) | 0.0793 |
Extinction coefficient (cm -1 M -1) | 2500001 |
Excitation (nm) | 656 |
Emission (nm) | 670 |
Quantum yield | 0.251 |
Storage, safety and handling
Certificate of Origin | Download PDF |
H-phrase | H301, H311, H331 |
Hazard symbol | T |
Intended use | Research Use Only (RUO) |
R-phrase | R23, R24, R25 |
Storage | Freeze (< -15 °C); Minimize light exposure |
UNSPSC | 12352200 |
Overview | SDSProtocol |
Molecular weight 1408.65 | Correction Factor (260 nm) 0.03 | Correction Factor (280 nm) 0.03 | Correction Factor (656 nm) 0.0793 | Extinction coefficient (cm -1 M -1) 2500001 | Excitation (nm) 656 | Emission (nm) 670 | Quantum yield 0.251 |
This deep red fluorescent phalloidin conjugate (equivalent to Alexa Fluor® 647-labeled phalloidin) selectively binds to F-actins. Used at nanomolar concentrations, phalloidin derivatives are convenient probes for labeling, identifying and quantitating F-actins in formaldehyde-fixed and permeabilized tissue sections, cell cultures or cell-free experiments. Phalloidin binds to actin filaments much more tightly than to actin monomers, leading to a decrease in the rate constant for the dissociation of actin subunits from filament ends, essentially stabilizing actin filaments through the prevention of filament depolymerization. Moreover, phalloidin is found to inhibit the ATP hydrolysis activity of F-actin. Phalloidin functions differently at various concentrations in cells. When introduced into the cytoplasm at low concentrations, phalloidin recruits the less polymerized forms of cytoplasmic actin as well as filamin into stable "islands" of aggregated actin polymers, yet it does not interfere with stress fibers, i.e. thick bundles of microfilaments. The property of phalloidin is a useful tool for investigating the distribution of F-actin in cells by labeling phalloidin with fluorescent analogs and using them to stain actin filaments for light microscopy. Fluorescent derivatives of phalloidin have turned out to be enormously useful in localizing actin filaments in living or fixed cells as well as for visualizing individual actin filaments in vitro. Fluorescent phalloidin derivatives have been used as an important tool in the study of actin networks at high resolution. AAT Bioquest offers a variety of fluorescent phalloidin derivatives with different colors for multicolor imaging applications.
Example protocol
AT A GLANCE
Protocol Summary
- Prepare samples in microplate wells
- Remove liquid from samples in the plate
- Add Phalloidin-iFluor™ 647 Conjugate solution (100 μL/well)
- Stain the cells at room temperature for 20 to 90 minutes
- Wash the cells
- Examine the specimen under microscope with Cy5 filter
Storage and Handling Conditions
The solution should be stable for at least 6 months if store at -20 °C. Protect the fluorescent conjugates from light, and avoid freeze/thaw cycles.Note Phalloidin is toxic, although the amount of toxin present in a vial could be lethal only to a mosquito (LD50 of phalloidin = 2 mg/kg), it should be handled with care.
PREPARATION OF STOCK SOLUTIONS
Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles.
Phalloidin-iFluor™ 647 Conjugate stock solution
Add 30 µL of DMSO into the powder and mix well.PREPARATION OF WORKING SOLUTION
Phalloidin-iFluor™ 647 Conjugate working solution
Add 1 µL of Phalloidin-iFluor™ 647 Conjugate solution to 1 mL of PBS with 1% BSA.Note The stock solution of phalloidin conjugate should be aliquoted and stored at -20 °C. protected from light.
Note Different cell types might be stained differently. The concentration of phalloidin conjugate working solution should be prepared accordingly.
SAMPLE EXPERIMENTAL PROTOCOL
Stain the cells
- Perform formaldehyde fixation. Incubate cells with 3.0–4.0 % formaldehyde in PBS at room temperature for 10–30 minutes.
Note Avoid any methanol containing fixatives since methanol can disrupt actin during the fixation process. The preferred fixative is methanol-free formaldehyde. - Rinse the fixed cells 2–3 times in PBS.
- Optional: Add 0.1% Triton X-100 in PBS into fixed cells for 3 to 5 minutes to increase permeability. Rinse the cells 2–3 times in PBS.
- Add 100 μL/well (96-well plate) of Phalloidin-iFluor™ 647 Conjugate working solution into the fixed cells, and stain the cells at room temperature for 20 to 90 minutes.
- Rinse cells gently with PBS 2 to 3 times to remove excess phalloidin conjugate before plating, sealing and imaging under microscope with Cy5 filter set.
Calculators
Common stock solution preparation
Table 1. Volume of DMSO needed to reconstitute specific mass of Phalloidin-iFluor® 647 Conjugate to given concentration. Note that volume is only for preparing stock solution. Refer to sample experimental protocol for appropriate experimental/physiological buffers.
0.1 mg | 0.5 mg | 1 mg | 5 mg | 10 mg | |
1 mM | 70.99 µL | 354.95 µL | 709.9 µL | 3.549 mL | 7.099 mL |
5 mM | 14.198 µL | 70.99 µL | 141.98 µL | 709.9 µL | 1.42 mL |
10 mM | 7.099 µL | 35.495 µL | 70.99 µL | 354.95 µL | 709.9 µL |
Molarity calculator
Enter any two values (mass, volume, concentration) to calculate the third.
Mass (Calculate) | Molecular weight | Volume (Calculate) | Concentration (Calculate) | Moles | ||||
/ | = | x | = |
Spectrum
Open in Advanced Spectrum Viewer
Spectral properties
Correction Factor (260 nm) | 0.03 |
Correction Factor (280 nm) | 0.03 |
Correction Factor (656 nm) | 0.0793 |
Extinction coefficient (cm -1 M -1) | 2500001 |
Excitation (nm) | 656 |
Emission (nm) | 670 |
Quantum yield | 0.251 |
Product Family
Name | Excitation (nm) | Emission (nm) | Extinction coefficient (cm -1 M -1) | Quantum yield | Correction Factor (260 nm) | Correction Factor (280 nm) |
Phalloidin-iFluor® 350 Conjugate | 345 | 450 | 200001 | 0.951 | 0.83 | 0.23 |
Phalloidin-iFluor® 405 Conjugate | 403 | 427 | 370001 | 0.911 | 0.48 | 0.77 |
Phalloidin-iFluor® 488 Conjugate | 491 | 516 | 750001 | 0.91 | 0.21 | 0.11 |
Phalloidin-iFluor® 514 Conjugate | 511 | 527 | 750001 | 0.831 | 0.265 | 0.116 |
Phalloidin-iFluor® 532 Conjugate | 537 | 560 | 900001 | 0.681 | 0.26 | 0.16 |
Phalloidin-iFluor® 555 Conjugate | 557 | 570 | 1000001 | 0.641 | 0.23 | 0.14 |
Phalloidin-iFluor® 594 Conjugate | 587 | 603 | 2000001 | 0.531 | 0.05 | 0.04 |
Phalloidin-iFluor® 633 Conjugate | 640 | 654 | 2500001 | 0.291 | 0.062 | 0.044 |
Phalloidin-iFluor® 680 Conjugate | 684 | 701 | 2200001 | 0.231 | 0.097 | 0.094 |
Show More (5) |
Images
Figure 1. Fluorescence images of HeLa cells stained with Phalloidin-iFluor® 647 Conjugate using fluorescence microscope with a Cy5 filter set (Red). Live cells were first stained with mitochondria dye MitoLite™ Green. After fixation in 4% formaldehyde, cells were labeled with Phalloidin-iFluor® 647 and counterstained with Nuclear Blue™ DCS1 (Cat#17548, Blue).
Figure 2. VG and IF images of the ORL. The periosteal (row A), intramuscular (row B), preorbicularis (row C) and dermal (row D) regions of the ORL were observed by VG (column 1) and IF (columns 2–5). There were immunopositive reactions for elastin (column 2, blue), collagen type I (column 3, green) and actin (column 4, red). The column 5 shows a merged image of the images in columns 2–4 image. Asterisks indicate confluence of the perimysium into the ORL fibres. Source: Three-dimensional structure of the orbicularis retaining ligament: an anatomical study using micro-computed tomography by Jehoon O et al., Scientific Reports, Nov. 2018.
Figure 3. Overall structure of the orbicularis retaining ligament (ORL). (a) Three-dimensional (3D) morphology reconstructed from micro-computed tomography (mCT) image sections. (b) Modified Verhoeff Van Gieson staining (VG) image. (c) A merged immunofluorescence (IF) image (elastin, blue; collagen type I, green; actin, red). Arrowheads indicate a direct fibre from the periosteum (P) to the dermis (D). OOc, orbicularis oculi muscle. S, sagittal; M, medial; A, anterior. Source: Three-dimensional structure of the orbicularis retaining ligament: an anatomical study using micro-computed tomography by Jehoon O et al., Scientific Reports, Nov. 2018.
Figure 5. Fluorescence images of HeLa cells stained with Phalloidin-iFluor® 647 Conjugate using fluorescence microscope with a Cy5 filter set (Red). HeLa cells were fixed with 4% formaldehyde followed by incubation with 1 ug/mL mouse tubulin antibody. Cells were stained with 10 ug/mL of GxM IgG- iFluor 488 conjugates. Cells were stained with Phalloidin-iFluor® 647 conjugate following product protocol and incubated with 2 uM DAPI for 5 min before imaging.
Figure 6. Co-localization of caveolin-1 and actin in post-conditioned hearts treated with latrunculin A. Representative confocal laser microscopy of the sections of frozen heart tissue after double-immunofluorescence staining of caveolin-1 (green) and actin (red). Cell nuclei were counterstained with DAPI. Source: Actin-Cytoskeleton Drives Caveolae Signaling to Mitochondria during Postconditioning by Correa et. al., Cells. Feb. 2023.
Figure 7. Cytokines induce intracellular component and cytoskeleton changes in RA synoviocytes. Holographic and tomographic microscopy images and phalloidin staining. Phalloidin-iFluor 647 was used for staining. Source: Effects of pro-inflammatory cytokines and cell interactions on cell area and cytoskeleton of rheumatoid arthritis synoviocytes and immune cells by Filali et. al., European Journal of Cell Biology. March 2023
Citations
View all 125 citations: Citation Explorer
Overstretching alveolar epithelial type II cells decreases surfactant secretion via actin polymerization and intracellular trafficking alteration
Authors: Inoue, Shigesato and Nagao, Junpei and Kawamoto, Kouhei and Kan-o, Keiko and Fukuyama, Satoru and Sasaki, Saori and Kudo, Susumu and Okamoto, Isamu and Sera, Toshihiro
Journal: Heliyon (2024)
Authors: Inoue, Shigesato and Nagao, Junpei and Kawamoto, Kouhei and Kan-o, Keiko and Fukuyama, Satoru and Sasaki, Saori and Kudo, Susumu and Okamoto, Isamu and Sera, Toshihiro
Journal: Heliyon (2024)
Sequential Fabrication of a Three-Layer Retina-Like Structure
Authors: Shechter, Yahel and Cohen, Roni and Namestnikov, Michael and Shapira, Assaf and Barak, Adiel and Barzelay, Aya and Dvir, Tal
Journal: Gels (2024): 336
Authors: Shechter, Yahel and Cohen, Roni and Namestnikov, Michael and Shapira, Assaf and Barak, Adiel and Barzelay, Aya and Dvir, Tal
Journal: Gels (2024): 336
Gelatin-Mediated Vascular Self-Assembly via a YAP-MMP Signaling Axis
Authors: Keshavarz, Mozhgan and Smith, Quinton
Journal: Advanced Functional Materials (2024): 2402360
Authors: Keshavarz, Mozhgan and Smith, Quinton
Journal: Advanced Functional Materials (2024): 2402360
Metal-organic frameworks as an active substrate for cell-interaction studies and cell-on-a-chip platforms
Authors: Jiang, Huijie and Gao, Ziyu and Lubrano, Claudia and Bovio, Claudia Latte and Bommes, Henning and Kauth, Andrea and Baumann, Lea and Cheng, Bo and Murugan, Divagar and Knoch, Joachim and others,
Journal: Biosensors and Bioelectronics: X (2024): 100487
Authors: Jiang, Huijie and Gao, Ziyu and Lubrano, Claudia and Bovio, Claudia Latte and Bommes, Henning and Kauth, Andrea and Baumann, Lea and Cheng, Bo and Murugan, Divagar and Knoch, Joachim and others,
Journal: Biosensors and Bioelectronics: X (2024): 100487
Programming Spatial Cell Sorting by Engineering Cadherin Intracellular Activity
Authors: Baba, Hikari and Fujita, Tomohiro and Mizuno, Kosuke and Tambo, Mai and Toda, Satoshi
Journal: ACS Synthetic Biology (2024)
Authors: Baba, Hikari and Fujita, Tomohiro and Mizuno, Kosuke and Tambo, Mai and Toda, Satoshi
Journal: ACS Synthetic Biology (2024)
Bioprinting of human dermal microtissues precursors as building blocks for endogenous in vitro connective tissue manufacturing
Authors: Scalzone, Annachiara and Imparato, Giorgia and Urciuolo, Francesco and Netti, Paolo Antonio
Journal: Biofabrication (2024)
Authors: Scalzone, Annachiara and Imparato, Giorgia and Urciuolo, Francesco and Netti, Paolo Antonio
Journal: Biofabrication (2024)
Microbiota-derived acetylcholine can promote gut motility in Drosophila melanogaster
Authors: Fujita, Yuka and Kosakamoto, Hina and Obata, Fumiaki
Journal: Philosophical Transactions of the Royal Society B (2024): 20230075
Authors: Fujita, Yuka and Kosakamoto, Hina and Obata, Fumiaki
Journal: Philosophical Transactions of the Royal Society B (2024): 20230075
2.5 D Actuating Substrates Enable Decoupling the Mechanical and Biochemical Effects of Muscle Exercise on Motor Neurons
Authors: Bu, Angel and Afghah, Ferdows and Castro, Nicolas and Bawa, Maheera and Kohli, Sonika and Shah, Karina and Rios, Brandon and Butty, Vincent and Raman, Ritu
Journal: bioRxiv (2024): 2024--03
Authors: Bu, Angel and Afghah, Ferdows and Castro, Nicolas and Bawa, Maheera and Kohli, Sonika and Shah, Karina and Rios, Brandon and Butty, Vincent and Raman, Ritu
Journal: bioRxiv (2024): 2024--03
TREM2 deficiency impairs the energy metabolism of Schwann cells and exacerbates peripheral neurological deficits
Authors: Zhang, Nannan and Ji, Qingjie and Chen, Yunfeng and Wen, Xiwu and Shan, Fangzhen
Journal: Cell Death \& Disease (2024): 193
Authors: Zhang, Nannan and Ji, Qingjie and Chen, Yunfeng and Wen, Xiwu and Shan, Fangzhen
Journal: Cell Death \& Disease (2024): 193
Modeling embryo-endometrial interface recapitulating human embryo implantation
Authors: Shibata, Shun and Endo, Shun and Nagai, Luis AE and H. Kobayashi, Eri and Oike, Akira and Kobayashi, Norio and Kitamura, Akane and Hori, Takeshi and Nashimoto, Yuji and Nakato, Ryuichiro and others,
Journal: Science Advances (2024): eadi4819
Authors: Shibata, Shun and Endo, Shun and Nagai, Luis AE and H. Kobayashi, Eri and Oike, Akira and Kobayashi, Norio and Kitamura, Akane and Hori, Takeshi and Nashimoto, Yuji and Nakato, Ryuichiro and others,
Journal: Science Advances (2024): eadi4819
References
View all 127 references: Citation Explorer
Improved penile histology by phalloidin stain: circular and longitudinal cavernous smooth muscles, dual-endothelium arteries, and erectile dysfunction-associated changes
Authors: Lin G, Qiu X, F and el TM, Albersen M, Wang Z, Lue TF, Lin CS.
Journal: Urology (2011): 970 e1
Authors: Lin G, Qiu X, F and el TM, Albersen M, Wang Z, Lue TF, Lin CS.
Journal: Urology (2011): 970 e1
Phalloidin perturbs the interaction of human non-muscle myosin isoforms 2A and 2C1 with F-actin
Authors: Diensthuber RP, Muller M, Heissler SM, Taft MH, Chizhov I, Manstein DJ.
Journal: FEBS Lett (2011): 767
Authors: Diensthuber RP, Muller M, Heissler SM, Taft MH, Chizhov I, Manstein DJ.
Journal: FEBS Lett (2011): 767
pH-(low)-insertion-peptide (pHLIP) translocation of membrane impermeable phalloidin toxin inhibits cancer cell proliferation
Authors: An M, Wijesinghe D, Andreev OA, Reshetnyak YK, Engelman DM.
Journal: Proc Natl Acad Sci U S A (2010): 20246
Authors: An M, Wijesinghe D, Andreev OA, Reshetnyak YK, Engelman DM.
Journal: Proc Natl Acad Sci U S A (2010): 20246
Labeling cytoskeletal F-actin with rhodamine phalloidin or fluorescein phalloidin for imaging
Authors: Chazotte B., undefined
Journal: Cold Spring Harb Protoc (2010): pdb prot4947
Authors: Chazotte B., undefined
Journal: Cold Spring Harb Protoc (2010): pdb prot4947
Protective effect of bile acid derivatives in phalloidin-induced rat liver toxicity
Authors: Herraez E, Macias RI, Vazquez-Tato J, Hierro C, Monte MJ, Marin JJ.
Journal: Toxicol Appl Pharmacol (2009): 21
Authors: Herraez E, Macias RI, Vazquez-Tato J, Hierro C, Monte MJ, Marin JJ.
Journal: Toxicol Appl Pharmacol (2009): 21
Anti-acetylated tubulin antibody staining and phalloidin staining in the starlet sea anemone Nematostella vectensis
Authors: Genikhovich G, Technau U.
Journal: Cold Spring Harb Protoc (2009): pdb prot5283
Authors: Genikhovich G, Technau U.
Journal: Cold Spring Harb Protoc (2009): pdb prot5283
Effect of Phalloidin on Filaments Polymerized from Heart Muscle Adp-Actin Monomers
Authors: Vig A, Dudas R, Kupi T, Orban J, Hild G, Lorinczy D, Nyitrai M.
Journal: J Therm Anal Calorim (2009): 721
Authors: Vig A, Dudas R, Kupi T, Orban J, Hild G, Lorinczy D, Nyitrai M.
Journal: J Therm Anal Calorim (2009): 721
In vitro inhibition of OATP-mediated uptake of phalloidin using bile acid derivatives
Authors: Herraez E, Macias RI, Vazquez-Tato J, Vicens M, Monte MJ, Marin JJ.
Journal: Toxicol Appl Pharmacol (2009): 13
Authors: Herraez E, Macias RI, Vazquez-Tato J, Vicens M, Monte MJ, Marin JJ.
Journal: Toxicol Appl Pharmacol (2009): 13
Processing of the phalloidin proprotein by prolyl oligopeptidase from the mushroom Conocybe albipes
Authors: Luo H, Hallen-Adams HE, Walton JD.
Journal: J Biol Chem (2009): 18070
Authors: Luo H, Hallen-Adams HE, Walton JD.
Journal: J Biol Chem (2009): 18070
Pygmy squids and giant brains: mapping the complex cephalopod CNS by phalloidin staining of vibratome sections and whole-mount preparations
Authors: Wollesen T, Loesel R, Wanninger A.
Journal: J Neurosci Methods (2009): 63
Authors: Wollesen T, Loesel R, Wanninger A.
Journal: J Neurosci Methods (2009): 63
Application notes
A Meta-Analysis of Common Calcium Indicators
A New Protein Crosslinking Method for Labeling and Modifying Antibodies
A Novel Fluorescent Probe for Imaging and Detecting Hydroxyl Radical in Living Cells
A Novel NO Wash Probeniceid-Free Calcium Assay for Functional Analysis of GPCR and Calcium Channel Targets
Abbreviation of Common Chemical Compounds Related to Peptides
A New Protein Crosslinking Method for Labeling and Modifying Antibodies
A Novel Fluorescent Probe for Imaging and Detecting Hydroxyl Radical in Living Cells
A Novel NO Wash Probeniceid-Free Calcium Assay for Functional Analysis of GPCR and Calcium Channel Targets
Abbreviation of Common Chemical Compounds Related to Peptides
FAQ
Are Cell Navigator® Cell Plasma Membrane Staining Kits suitable for cell culture medium samples?
Are there any alternatives to BrdU (Bromodeoxyuridine)?
Are there any alternatives to Cy5?
Are there any alternatives to indocyanine green (ICG)?
Are there any calcium indicators that don't require probenecid (PBC)?
Are there any alternatives to BrdU (Bromodeoxyuridine)?
Are there any alternatives to Cy5?
Are there any alternatives to indocyanine green (ICG)?
Are there any calcium indicators that don't require probenecid (PBC)?