Revision 1

#8606Store at -20C

1 Kit

(7 x 20 microliters)

Cell Signaling Technology

Orders: 877-616-CELL (2355) [email protected]

Support: 877-678-TECH (8324)

Web: [email protected] cellsignal.com

3 Trask LaneDanversMassachusetts01923USA
For Research Use Only. Not for Use in Diagnostic Procedures.
Product Includes Product # Quantity Mol. Wt Isotype/Source
Phospho-Rac1/cdc42 (Ser71) Antibody 2461 20 µl 28 kDa Rabbit 
Rac1/Cdc42 Antibody 4651 20 µl 21 kDa Rabbit 
WAVE-2 (D2C8) XP® Rabbit mAb 3659 20 µl 80 kDa Rabbit IgG
Profilin-1 (C56B8) Rabbit mAb 3246 20 µl 15 kDa Rabbit IgG
ARP2 Antibody 3128 20 µl 44 kDa Rabbit 
ARP3 Antibody 4738 20 µl 47 kDa Rabbit 
N-WASP (30D10) Rabbit mAb 4848 20 µl 65 kDa Rabbit IgG
Anti-rabbit IgG, HRP-linked Antibody 7074 100 µl Goat 

Please visit cellsignal.com for individual component applications, species cross-reactivity, dilutions, protocols, and additional product information.

Description

The Actin Nucleation and Polymerization Antibody Sampler Kit provides an economical means to evaluate the presence and status of actin nucleation and polymerization. The kit contains enough primary antibody to perform two western blots per primary.

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Background

Actin nucleation is the process of forming new actin filaments and is necessary to stimulate actin polymerization. Actin polymerization is vital for cell motility, cell division, and cell adhesion. Rac and Cdc42, members of the Rho-GTPase family, play key roles in actin dynamics, membrane trafficking, transcriptional regulation, cell growth, and development (1). GTP binding stimulates the activity of Rac/Cdc42, and the hydrolysis of GTP to GDP through the intrinsic GTPase activity or Rac/Cdc42, rendering it inactive. GTP hydrolysis is aided by GTPase activating proteins (GAPs), while exchange of GDP for GTP is facilitated by guanine nucleotide exchange factors (GEFs). Another level of regulation is achieved through binding of RhoGDI, a guanine dissociation inhibitor, which retains Rho family GTPases, including Rac and Cdc42, in their inactive GDP-bound state (2,3). Hematopoietic WASP and ubiquitously expressed N-WASP are autoinhibited in unstimulated cells. Upon stimulation they are activated by Cdc42, which relieves the autoinhibition in conjunction with phosphatidyl inositol 4,5-bisphosphate (4). Three WAVE (Wasf, SCAR) family proteins are similar in sequence to WASP and N-WASP, but lack the WASP/N-WASP autoinhibition domains and are indirectly activated by Rac (4). WAVE-2 is widely distributed, while WAVE-1 and WAVE-3 are strongly expressed in the brain (5). The highly conserved ARP2/3 complex is an important actin nucleation protein complex consisting of ARP2, ARP3, and ARPC1-5. The ARP2/3 complex promotes branching of existing actin filaments and formation of daughter filaments following activation by nucleation-promoting factors, such as WASP/WAVE or cortactin (6). Profilins are conserved actin binding proteins that affect the rate of actin polymerization by binding actin monomers and promoting exchange of ADP for ATP (reviewed in 7). Profilins bind to proteins involved in the regulation of actin dynamics including paladin (8), dynamin-I (9), VASP (10), and N-WASP (11).

  1. Wennerberg, K. and Der, C.J. (2004) J Cell Sci 117, 1301-12.
  2. Bernards, A. and Settleman, J. (2004) Trends Cell Biol 14, 377-85.
  3. Rossman, K.L. et al. (2005) Nat Rev Mol Cell Biol 6, 167-80.
  4. Millard, T.H. et al. (2004) Biochem J 380, 1-17.
  5. Suetsugu, S. et al. (1999) Biochem Biophys Res Commun 260, 296-302.
  6. Goley, E.D. and Welch, M.D. (2006) Nat Rev Mol Cell Biol 7, 713-26.
  7. Witke, W. (2004) Trends Cell Biol 14, 461-9.
  8. Boukhelifa, M. et al. (2006) FEBS J 273, 26-33.
  9. Gareus, R. et al. (2006) J Biol Chem 281, 2803-11.
  10. Reinhard, M. et al. (1995) EMBO J 14, 1583-9.
  11. Suetsugu, S. et al. (1998) EMBO J 17, 6516-26.

Background References

    Trademarks and Patents

    Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
    U.S. Patent No. 7,429,487, foreign equivalents, and child patents deriving therefrom.
    All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

    Limited Uses

    Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

    Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

    Revision 1
    #8606

    Actin Nucleation and Polymerization Antibody Sampler Kit

    Actin Nucleation and Polymerization Antibody Sampler Kit: Image 1 Expand Image
    Western blot analysis of A431 cell extracts, untreated (-) or EGF treated (+), using Phospho-Rac1/cdc42 (Ser71) Antibody (upper) or Rac1/Cdc42 Antibody #4651 (lower).
    Actin Nucleation and Polymerization Antibody Sampler Kit: Image 2 Expand Image
    Western blot analysis of extracts from various cell types using ARP2 Antibody.
    Actin Nucleation and Polymerization Antibody Sampler Kit: Image 3 Expand Image
    Western blot analysis of extracts from various cell types using Profilin-1 (C56B8) Rabbit mAb.
    Actin Nucleation and Polymerization Antibody Sampler Kit: Image 4 Expand Image
    Western blot analysis of extracts from various cell types using WAVE-2 (D2C8) XP® Rabbit mAb.
    Actin Nucleation and Polymerization Antibody Sampler Kit: Image 5 Expand Image
    Western blot analysis of extracts from various cell lines using Rac1/Cdc42 Antibody.
    Actin Nucleation and Polymerization Antibody Sampler Kit: Image 6 Expand Image
    Western blot analysis of extracts from HeLa, Jurkat and COS cells using ARP3 Antibody.
    Actin Nucleation and Polymerization Antibody Sampler Kit: Image 7 Expand Image
    Western blot analysis of extracts from various cell types, using N-WASP (30D10) Rabbit mAb.
    Actin Nucleation and Polymerization Antibody Sampler Kit: Image 8 Expand Image
    After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.
    Actin Nucleation and Polymerization Antibody Sampler Kit: Image 9 Expand Image
    Immunoprecipitation of WAVE-2 from HeLa cells using WAVE-2 (D2C8) XP® Rabbit mAb. Western blot was performed using the same antibody. Lane 1 is 5% input.
    Actin Nucleation and Polymerization Antibody Sampler Kit: Image 10 Expand Image
    Immunoprecipitation of N-WASP from MCF-7 cells, using N-WASP (30D10) Rabbit mAb. Western blot was performed using the same antibody.
    Actin Nucleation and Polymerization Antibody Sampler Kit: Image 11 Expand Image
    Confocal immunofluorescent analysis of HeLa cells, serum-starved (left) or EGF-treated #9908 (right), using Wave-2 (D2C8) XP® Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).