Overview
Alternative Name Neurotoxin 2, Toxin ATX-II, As2, Anemonia viridis toxin 2, Av2, Delta-actitoxin-Avd1c 2, Toxin II
Lyophilized Powder yes
Origin Anemonia sulcata (Mediterranean snakelocks sea anemone)
Source Synthetic modified peptide
MW: 5561 Da
Form Lyophilized from double distilled water (ddH2O). May contain TFA as a residual counter ion.
Effective concentration 0.1 – 1 µM
Sequence GVPCLCDSDGPSVRGNTLSGIIWLAGCPSGWHNCKKHGPTIGWCCKQ-OH
Modifications Disulfide bonds between: Cys4-Cys44, Cys6-Cys34 and Cys27-Cys45
ATTO Fluor-647N
ATTO Fluor-647N
Label ATTO-647N. ATTO dyes are characterized by strong absorption (high extinction coefficient), high fluorescence quantum yield, and high photo-stability. Maximum absorption 646 nm; maximum fluorescence 664 nm. The fluorescence is excited most efficiently in the range 625 - 660 nm. A suitable excitation source is the 647 nm line of the Krypton-Ion laser or a diode-laser emitting at 650 nm. It can be used in flow cytometry (FACS) using the red (637 nm) laser. The extent of labeling is 1-2 molecules of dye per ATX-II molecule.
Structure
Activity ATX-II potently modulates voltage-gated Na+ channel gating kinetics by delaying its inactivation and prolonging the action potential of excitable membranes. ATX-II has been used as a powerful activator of TTX-sensitive and insensitive Na+ channels in various excitable tissue and cell types1-9.
References-Activity
- Romey, G. et al. (1976) Proc. Natl. Acad. U.S.A. 73, 4055.
- Tesseraux, I. et al. (1987) Naunyn Schmiedebergs Arch. Pharmacol. 336, 232.
- Nishio, M. et al. (1991) Br. J. Pharmacol. 104, 504.
- Cannon, S.C. and Corey, D.P. (1993) J. Physiol. 466, 501.
- Hoey, A. et al. (1994) Pharmacol. Toxicol. 75, 356.
- Chahine, M. et al. (1996) J. Membr. Biol. 152, 39.
- Chahine, M. et al. (1996) J. Membr. Biol. 152, 39.
- Brand, S. et al. (2000) Eur. J. Neurosci. 12, 2387.
- Ravens, U. (1976) Naunyn Schmiedebergs Arch. Pharmacol. 296, 73.
Accession number P30288
Shipping and storage The product is shipped as a lyophilized powder at room temperature. Upon receipt, store the product at -20°C. Protect from moisture. Avoid exposure to light.
Solubility The product is lyophilized in 0.5 ml conical vial.
Centrifuge the vial before adding solvent (10,000 x g for 5 minutes). The lyophilizate may be difficult to visualize. Add solvent directly to the centrifuged vial. Tap the vial to aid in dissolving the lyophilized product. Tilt and gently roll the liquid over the walls of the vial. Avoid vigorous vortexing. Light vortexing for up to 3 seconds is acceptable if needed.
Soluble in pure water at high-micromolar concentrations (50 µM - 1 mM). For long-term storage in solution, it is recommended to prepare a stock solution by dissolving the product in double distilled water (ddH2O) at a concentration between 100-1000x of the final working concentration. Divide the stock solution into small aliquots and store at -20°C. Before use, thaw the relevant vial(s) and dilute to the desired working concentration in your working buffer. Centrifuge all product preparations at 10,000 x g for 5 minutes before use. It is recommended to prepare fresh solutions in working buffers just before use. Avoid multiple freeze-thaw cycles to maintain biological activity. Avoid exposure to light.
Centrifuge the vial before adding solvent (10,000 x g for 5 minutes). The lyophilizate may be difficult to visualize. Add solvent directly to the centrifuged vial. Tap the vial to aid in dissolving the lyophilized product. Tilt and gently roll the liquid over the walls of the vial. Avoid vigorous vortexing. Light vortexing for up to 3 seconds is acceptable if needed.
Soluble in pure water at high-micromolar concentrations (50 µM - 1 mM). For long-term storage in solution, it is recommended to prepare a stock solution by dissolving the product in double distilled water (ddH2O) at a concentration between 100-1000x of the final working concentration. Divide the stock solution into small aliquots and store at -20°C. Before use, thaw the relevant vial(s) and dilute to the desired working concentration in your working buffer. Centrifuge all product preparations at 10,000 x g for 5 minutes before use. It is recommended to prepare fresh solutions in working buffers just before use. Avoid multiple freeze-thaw cycles to maintain biological activity. Avoid exposure to light.
Storage of solutions Store the reconstituted solution at -20°C for the shortest time possible. Avoid multiple freeze-thaw cycles. We do not recommend storing the product in working solutions for longer than a day. Avoid exposure to light.
Our bioassay
Live cell imaging of ATX-II-ATTO Fluor-647N in neonatal mouse cardiomyocytes.Neonatal mouse cardiomyocytes were isolated from hearts of two days old neonatal mice. (A) CellMask™ Actin 1X solution was applied for 30 minutes, resulting in a green fluorescence to visualize cellular membrane. (B) Following this, the same cells underwent incubation with 0.3 µM of ATX-II-ATTO Fluor-647N (#STA-700-FRN) for 60 minutes at 37ºC, followed by PBSX1 wash, leading to red fluorescence indicative of the distribution of Na+ channels. (C) Live imaging of the cardiomyocytes allowed observation of ATX-II distribution among the cells.
Unlabeled ATX-II successfully blocks Na+ channels access.Neonatal mouse cardiomyocytes were isolated from hearts of two days old neonatal mice. (A) CellMask™ Actin 1X solution was applied for 30 minutes, resulting in a green fluorescence to visualize cellular membrane. (B) Following this, the same cells underwent incubation with 50 µM of ATX-II (#STA-700), and 0.3 µM of ATX-II-ATTO Fluor-647N (#STA-700-FRN) for 60 minutes at 37ºC, followed by PBSX1 wash. (C) Live imaging of cardiomyocytes cells demonstrates that ATX-II successfully competes with ATX-II-ATTO Fluor-647N for binding sites of Na+ channels.
ATX-II co-localizes with NaV1.5 (SCN5A) in mouse heart, L&R ventricle frozen sections.(A) Frozen sections underwent incubation with Anti-NaV1.5 (extracellular)-ATTO Fluor-550 Antibody (ASC-045-AO, 1:50) leading to orange fluorescence. (B) The same frozen sections underwent incubation with 0.3 µM ATX-II-ATTO Fluor-647N (#STA-700-FRN), leading to red fluorescence. (C-D) Following this, CellMask™ Actin 1X solution and Hoechst 33342 (2 µg/ml) was applied for 15 minutes, resulting in a green and blue fluorescence to visualize cellular membrane and nuclei, respectively. (E) Imaging of heart frozen sections allowed observation of NaV1.5 and ATX-II co-localization.
Unlabeled ATX-II successfully blocks access to NaV1.5 (SCN5A) in mouse heart, L&R ventricle frozen sections.(A) Frozen sections underwent incubation with Anti-NaV1.5 (extracellular)-ATTO Fluor-550 Antibody (ASC-045-AO, 1:50) leading to orange fluorescence. (B) The same frozen sections underwent incubation with 0.3 µM ATX-II-ATTO Fluor-647N (#STA-700-FRN) and 100 µM ATX-II (#STA-700). (C-D) Following this, CellMask™ Actin 1X solution and Hoechst 33342 (2 µg/ml) was applied for 15 minutes, resulting in a green and blue fluorescence to visualize cellular membrane and nuclei, respectively. (E) Imaging of frozen sections demonstrates that ATX-II successfully competes with ATX-II-ATTO Fluor-647N for binding sites of NaV1.5.
Direct flow cytometry of ATX-II in live intact human AC16 cells.___ AC16 cells.
___ AC16 cells + 0.1 µM ATX-II (#STA-700).
___ AC16 cells + 0.1 µM ATX-II-ATTO Fluor-647N (#STA-700-FRN).
Direct flow cytometry of ATX-II in live intact human HEK 293T Nav1.5 cells.___ Nav1.5 cells.
___ Nav1.5 cells + 0.1 µM ATX-II (#STA-700).
___ Nav1.5 cells + 0.1 µM ATX-II-ATTO Fluor 647N (#STA-700-FRN).
Alomone Labs ATX-II-ATTO Fluor-647N enhances the current of NaV1.5 channels expressed in Xenopus oocytes.A. Representative time course of ATX-II-ATTO Fluor-647N (#STA-700-FRN) effect on the normalized area of NaV1.5 channels current. Membrane potential was held at -100 mV, current was elicited by a 100 ms voltage step to 0 mV every 10 sec, and was significantly enhanced by the application of 0.5 μM ATX-II-ATTO Fluor-647N (green).
B. Superimposed traces of NaV1.5 current after application of control (black) and of 0.5 μM ATX-II-ATTO Fluor-647N (green), taken from the recording in A.
References - Scientific background
- Romey, G. et al. (1976) Proc Natl. Acad. Sci. U.S.A. 73, 4055.
- Tesseraux, I. et al. (1987) Naunyn Schmiedebergs Arch. Pharmacol. 336, 232.
- Nishio, M. et al. (1991) Br. J. Pharmacol. 104, 504.
- Cannon, S.C. and Corey D.P. (1993) J. Physiol. 466, 501.
- Hoey, A. et al. (1994) Pharmacol. Toxicol. 75, 356.
- Chahine, M. et al. (1996) J. Membr. Biol. 152, 39.
- Fletcher, J.E. et al. (1999) Anesthesiology 90, 1294.
- Brand, S. et al. (2000) Eur. J. Neurosci. 12, 2387.
- Ravens, U. (1976) Naunyn-Schmiedeberg’s Arch. Pharmacol. 296, 73.
Scientific background
ATX-II is a 47 amino acid peptidyl toxin, originally isolated from Anemonia sulcata sea anemone venom. It is a potent neurotoxin, which modulates voltage-gated Na+ channel gating kinetics by delaying its inactivation and prolonging the action potential of excitable membranes.
ATX-II has been used as a powerful activator of TTX-sensitive and -insensitive Na+ channels in various excitable tissue and cell types (at concentration range of 10-100 nM)1-9.
Target Various NaV channels
Lyophilized Powder
ATX-II-ATTO Fluor-647N (STA-700-FRN) is a highly pure, synthetic, and biologically active conjugated toxin.
Benefits of ATX-II-ATTO Fluor-647N:
✓ Localization and distribution
✓ Clustering and internalization kinetics
✓ Live cell imaging
✓ Single cell detection
✓ Binding kinetics
✓ Direct flow cytometry
We gladly take on collaboration projects. Please Contact Us.
For research purposes only, not for human use
Last Update: 15/12/2024
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