Actin-Toolkit TIRFM
The complete solution!
ATTO565-labeled native α-actin from rabbit skeletal muscle
Cat. #: 8095-01
Applications
- Single-molecule analysis of actin polymerization, depolymerization and nucleation by TIRF microscopy.
- Visualization of ligand-induced changes in actin filament dynamics.
- Fluorescence-based analysis of actin filaments, bundles and networks under defined in vitro conditions.
Description
| Product name | Actin-Toolkit TIRFM | ATTO565-Actin |
| Protein | ATTO565-Actin (α-actin) |
| Origin | Rabbit skeletal muscle |
| Molecular mass | 42 kDa |
| Toolkit description |
The Actin-Toolkit TIRFM is designed for direct visualization of actin dynamics by total internal reflection fluorescence microscopy and related fluorescence-based single-molecule imaging approaches. The toolkit contains ATTO565-labeled rabbit skeletal muscle α-actin with a Degree of Labeling (DOL) of 0.3. The labeled actin is intended for incorporation into actin filaments at a low labeling fraction suitable for fluorescence imaging of filament assembly and dynamics. NEM-Myosin II is supplied as a surface-tethering component for flow-chamber assays. The kit also contains passivation solution, actin polymerization and monomer buffers, Mg-exchange buffer and FluMaXx oxygen-scavenging reagents for fluorescence imaging under reduced photochemical stress. A dedicated handbook describes flow-chamber assembly and TIRFM actin-polymerization assays. |
| Degree of Labeling | DOL: 0.3 |
| Optical properties | ATTO565: λabs 563 nm, λfl 592 nm |
| Purity | >99% by scanning densitometry. |
Properties
| Form | Lyophilized and in solution, depending on kit component. |
| Kit content | 5 × 100 µg ATTO565-Actin (lyophilized powder); 1 × 100 µg NEM-Myosin II (lyophilized powder); 3 × 1 ml Passivation solution (5% BSA, fatty-acid free); 1 × 1 ml PolyMix (lyophilized F-actin ATP buffer); 1 × 50 ml MonoMix (lyophilized G-actin ATP buffer); 1 × 2 ml HSB, 5× stock (Myosin Buffer, pH 7.0); 1 × 0.5 ml ME-Buffer (100× Mg-exchange buffer); 1 × Toolkit Handbook TIRFM. |
| FluMaXx components |
5 × 250 µl FluMaXx (2× stock, lyophilized);
Contains glucose oxidase, catalase, imidazole pH 7.4, KCl, EGTA, MgCl2, glucose and cryoprotectants (proprietary formula); 1 × 500 µl ATP/DTT Mix (10× stock, lyophilized); 2 × 1.5 ml methylcellulose stock solution (0.5% in Milli-Q™ water). |
| Actin buffer after reconstitution | 2 mM Tris-Cl, pH 8.2, 0.4 mM ATP, 0.5 mM DTT, 0.1 mM CaCl2 and 0.3% disaccharides when reconstituted with 1.0 ml ultrapure water to obtain a 1 mg/ml actin solution. |
| Related products | Actin-binding proteins and additional actin reagents for defined in vitro reconstitution experiments. |
| Purification notes | Native actin purified from rabbit skeletal muscle; final purification by gel permeation chromatography (GPC). ATTO565 conjugate prepared from purified actin. |
| Protein concentration | Actin determined by direct UV absorbance at A290 nm; A290nm 1% = 0.62. |
| Storage instructions | We guarantee stable performance of the kit components for 6 months when stored at −70 °C upon arrival. Solubilized G-actin should be kept on ice and used within 1 week. Avoid refreezing. |
| Shipping conditions | Shipped at ambient temperature. Upon delivery store at −70 °C. |
| CAS no. | 51005-14-2 (Actin) |
| Remarks | For research use only. Not for diagnostic, therapeutic, clinical or in vivo use. |
Buffers and proteins from Hypermol® are made using ultrapure reagents in Milli-Q™ water, as described in our publications.
Further Information
References
Modulation of actin dynamics as potential macrophage subtype-targeting anti-tumour strategy.
Pergola C, Schubert K, Pace S, et al. Scientific Reports. 2017;7:41434. doi:10.1038/srep41434.
Actin-Toolkit TIRFM
The complete solution!
ATTO565-Actin, rabbit skeletal muscle actin
Cat. # 8095-01
.............................................................................................................................................................................................................................................................................................................................................................................................................
Applications
- Single molecule analysis of actin polymerization, depolymerization and nucleation;
- Visualization of ligand-induced actin dynamics;
- High- and superresolution analysis of actin molecules, filaments, bundles and networks;
Description
|
|
|
|
Protein |
ATTO565-Actin (α-Actin) |
|
Origin |
Skeletal muscle alpha actin, rabbit |
|
Molecular mass |
42kDa |
|
Toolkit description |
The Actin-Toolkit TIRFM was developed to analyze actin dynamics by TIRF microscopy or other fluorescence based single molecule imaging techniques. The Toolkit contains all components required for TIRFM analyses and enables the user to start TIRFM experiments straight ahead. It includes ATTO565-Actin (λex 563nm, λem 592nm) with a DOL of 0.3 as demanded for TIRFM analysis. The Toolkit contains NEM-Myosin II as a tethering protein, and all components required for flow chamber TIRF assays, including FluMaXx - an efficient oxygen scavenger system - to protect actin filament from fragmentation during analyses. A detailed description of the flow chamber assembly and TIRFM actin polymerization assays is provided with the handbook. In combination with the actin buffers contained in the kit, error-free handling is guaranteed. The kit contains 5x100 µg of α-skeletal muscle ATTO565-Actin conjugates and buffers to keep the actin functional. |
|
Purity |
>99% by scanning densitometry. |
Properties
|
|
|
|
Form |
Lyophilized and in solution (see kit content). |
|
Kit content |
5x100 µg ATTO565-Actin (lyophilized powder);
5x250 µl FluMaXx (2x stock, lyophilized) |
|
Actin Buffer |
2mM Tris-Cl pH 8.2, 0.4mM ATP, 0.5mM DTT, 0.1mM CaCl2, 1mM NaN3 and 0.3% disaccharides, when reconstituted with 1.0 ml ultrapure water to obtain a 1mg/ml solution. |
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Related products |
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Purification notes |
Purified from rabbit skeletal muscle, GPC. |
|
Protein concentration |
Determined by the Biuret method. |
|
Storage instructions |
We guarantee stable performance of the kit components for 6 months when stored at -70°C upon arrival. Solubilized G-actin is kept on ice and should be used within 1 week. Avoid refreezing. |
|
Shipping conditions |
At ambient temperature. Upon delivery store at -70°C. |
|
Remarks |
For Use in Research only. Not for Use in Human or Veterinary Diagnostical or Therapeutical Applications. |
|
CAS no. |
51005-14-2 (Actin) |
Further Information
Material and Safety Data Sheet
Table of Dyes and Light Sources 
Protein Sequence on NCBI
References
Modulation of actin dynamics as potential macrophage subtype-targeting anti-tumour strategy.
Pergola C, Schubert K, Pace S, Ziereisen J, Nikels F, Scherer O, Hüttel S, Zahler S, Vollmar AM, Weinigel C, Rummler S, Müller R, Raasch M, Mosig A, Koeberle A, Werz O.
Sci Rep. 2017 Jan 30;7:41434. doi: 10.1038/srep41434.
Handbook Actin-Toolkit TIRFM
(Web version only for demonstration purpose!
A print version comes with the kit)
Flyer Actin-Toolkits
Material and Safety Data Sheet
Protein Sequence on NCBI
