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Wave2

Manufactured by Cell Signaling Technology
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WAVE2 is a laboratory equipment product from Cell Signaling Technology. It is a key regulator of actin cytoskeleton reorganization and is involved in various cellular processes.

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Lab products found in correlation

P34arc, by Merck Group (3 mentions) α tubulin, by Cell Signaling Technology (2 mentions) Pan actin, by Cell Signaling Technology (2 mentions) N wasp, by Cell Signaling Technology (2 mentions) Hsc70 mab, by Santa Cruz Biotechnology (2 mentions) Myc mab, by Merck Group (2 mentions) Ptyr mab, by Merck Group (2 mentions) Gfp mab, by Roche (2 mentions) Alexa fluor 488 anti rabbit igg, by Thermo Fisher Scientific (1 mentions) Anti mouse igg alexa fluor 594, by Thermo Fisher Scientific (1 mentions) Alexa fluor 647 anti rat igg, by Thermo Fisher Scientific (1 mentions) E cadherin, by Cell Signaling Technology (1 mentions) β actin, by Cell Signaling Technology (1 mentions) Alexa fluor 488 anti mouse igg, by Thermo Fisher Scientific (1 mentions) Kinomeview profiling kit, by Cell Signaling Technology (1 mentions) Anti mouse ig hrp, by Agilent Technologies (1 mentions) Anti rabbit ig hrp, by Agilent Technologies (1 mentions) Vimentin, by Cell Signaling Technology (1 mentions) Ripa buffer, by Thermo Fisher Scientific (1 mentions) Chemidoc touch imaging system, by Bio-Rad (1 mentions)

Spelling variants of this product

Rabbit anti-WAVE2 (2 citations)

10 protocols using wave2

1

Antibody Validation Protocols for Western Blot and Immunofluorescence

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Antibodies were used for both Western blots and immunofluorescence. Primary antibodies for Arp2, Arp3, α-tubulin, β-actin, E-cadherin, KinomeView Profiling kit, N-WASP, pan-actin, phospho-PLK1 (Thr210), PLK1, phospho-Rac1/Cdc42(Ser71), Rac1/Cdc42, vimentin, and WAVE-2 were purchased from Cell Signaling Technology (#3128, #4738, #2144, #4970, #3195, #9812, #4848, #8456, #9062, #4513, #2461, #4651, #5741, and #3659). PLK1 primary antibody used for immunofluorescence was purchased from Thermo Fisher Scientific (#37-7100); γ-actin antibody was purchased from Abcam (#ab123034). HCV NS5A sheep polyclonal antibody and NS5A mouse antibody were gifts from Professor Mark Harris (University of Leeds, UK) and Associate Professor Michael Beard (University of Adelaide), respectively. Secondary antibodies were either conjugated with HRP; anti-rabbit Ig/HRP (#P0448), and anti-mouse Ig/HRP (#P0161) (DAKO) or labelled with fluorescence dyes: antirabbit IgG Alexa Fluor 488 (#A21206), anti-mouse IgG Alexa Fluor 488 (#A21202), anti-mouse IgG Alexa Fluor 594 (#A21203), and anti-rat IgG Alexa Fluor 647 (#A21472) (Thermo Fisher Scientific).
El-Khobar K.E., Tay E., Diefenbach E., Gloss B.S., George J, & Douglas M.W. (2023). Polo-like kinase-1 mediates hepatitis C virus-induced cell migration, a drug target for liver cancer. Life Science Alliance, 6(11), e202201630.
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2

Western Blot Analysis of Actin Regulators

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For cell lysate preparation, BMDMs transfected either with mimic control or miR-181c-5p mimic (in the presence or absence of apoptotic H9c2 cells) were lysed in RIPA buffer (Cat# J63324, Alfa Aesar) with protease and phosphatase inhibitor cocktail. Protein concentrations were determined by Bradford assay (Bio-Rad, USA). Equal amounts of proteins were denatured in 4× Laemmli buffer, separated on denaturing SDS-PAGE gel (4–20%), and then transferred to 0.2 μm PVDF membranes. The membranes were blocked with 5% bovine serum albumin (BSA) or non-fat milk powder (w/v) in TBS-T for 1 h at room temperature. The membranes were incubated with primary antibodies; N-WASP, WAVE-2 (actin nucleation and polymerization antibody sampler kit, Cat# 8606, Cell Signaling Technology, 1:1000 dilution for each antibody), and β-Tubulin (Proteintech, Cat# 66240-1-Ig, 1:10000) overnight at 4 °C, followed by HRP-conjugated secondary antibodies. The protein bands were developed by enhanced chemiluminescence (Pierce) detection system. Images of the blots were acquired on a ChemiDoc™ Touch Imaging System (Bio-Rad, USA), and densitometric analyses were performed using ImageJ (NIH) software.
Singh S., Henderson J.C., Patil M., Dubey P.K., Dubey S., Kannappan R., Zhang J, & Krishnamurthy P. (2022). MicroRNA-181c-5p modulates phagocytosis efficiency in bone marrow-derived macrophages. Inflammation research : official journal of the European Histamine Research Society ... [et al.], 71(3), 321-330.
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3

Western Blot Analysis of Protein Expression

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Cell or tissue lysates were resolved on sodium dodecyl sulfate–polyacrylamide gel electrophoresis gels and transferred to Nitrocellulose membrane (Axygen, CA, USA). After blocking with 5% non-fat milk, the membrane was incubated with primary antibodies against with c-Met (#1996-1, Epitomics, Burlingame, CA, USA), WAVE2 (#3659, Cell Signaling Technology, Danvers, MA, USA), Sox9 (#AB5535, Millipore, Billerica, MA, USA), Vinculin (#4650, Cell Signaling Technology), or β-actin (CP01; Calbiochem, San Diego, CA, USA) overnight at 4 °C and then incubated with horseradish peroxidase-conjugated secondary antibody. The signal was detected in a sensitive digital imaging equipment (ImageQuant LAS 4000 mini, GE Healthcare, Piscataway, NJ, USA) using the ECL detection kit (Millipore). The protein fragments were quantified by densitometry using Quantity One software (Bio-Rad).
Wang Q.Y., Zhou C.X., Zhan M.N., Tang J., Wang C.L., Ma C.N., He M., Chen G.Q., He J.R, & Zhao Q. (2018). MiR-133b targets Sox9 to control pathogenesis and metastasis of breast cancer. Cell Death & Disease, 9(7), 752.
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4

Comprehensive Protein Expression Analysis

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Trypsin-EDTA (Cat# 25-053-CI), PBS (Cat# 21-031-CV), and laminin (Cat# 354232) were purchased from Corning. Recombinant EGF was purchased from Life Technologies (Cat# PHG0311). IGF was purchased from Sigma-Aldrich (Cat# I3769). PIM447 was purchased from Selleck Chemicals (Cat# S7985). AZD1208 was acquired from AdooQ Bioscience (Cat# A13203-750). DMSO was purchased from Thermo Fisher Scientific (Cat# 97064-724). Cycloheximide was purchased from VWR (Cat# 97064-724), and doxycycline was purchased from Sigma-Aldrich (Cat# D9891-5G). Recombinant myelin basic protein (MBP) was a gift from Dr. Greg Rogers, and recombinant ABI2 protein was purchased from Origene (Cat# TP300637). Radio-labeled ATP was purchased from Perkin Elmer (Cat# BLU502A). The antibody to ABI2 was purchased from Bethyl Laboratories (Cat# A302-499A-M). GFP (Cat# 2956S), HA (Cat# 3724S), HIF-1a (Cat# 14179S), p-IRS1 [S1101] (Cat# 2385S), PIM1 (Cat# 3247S), and WAVE2 (Cat# 3659S) antibodies were purchased from Cell Signaling Technology. The antibody for actin was purchased from BD Biosciences (Cat# 61656). PIM1 (Cat# ab75776) and Ki67 (Cat#ab833) antibodies used for immunohistochemistry were purchased from Abcam.
Jensen C.C., Clements A.N., Liou H., Ball L.E., Bethard J.R., Langlais P.R., Toth R.K., Chauhan S.S., Casillas A.L., Daulat S.R., Kraft A.S., Cress A.E., Miranti C.K., Mouneimne G., Rogers G.C, & Warfel N.A. (2023). PIM1 phosphorylates ABI2 to enhance actin dynamics and promote tumor invasion. The Journal of Cell Biology, 222(6), e202208136.
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5

Visualizing Cytoskeletal Protein Localization

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Cells were plated on uncoated coverslips, and grown as indicated. Prior to imaging, cells were fixed 15 min in 3.7% paraformaldehyde, and permeabilized in 0.2% Triton X-100 for 5 min. Cells were incubated with either Phalloidin (1:200, to detect F-Actin) or primary antibodies to Cortactin (1:400, Upstate Biotech), WAVE2 (1:400, Cell Signaling) or Nck (1:400, BD Biosciences). AlexaFluor conjugated secondary antibodies (1:8000 to 1:10,000, Thermo Fisher Scientific) were then used to visualize protein localization. Cell images were captured using a Zeiss Axioplan 2 with 40X, NA 0.75 objective and Zeiss AxioCam CCD Camera. Pseudocolor images were created in Photoshop using green-magenta color scheme in which colocalization appears white.
Kadrmas J.L., Beckerle M.C, & Yoshigi M. (2020). Genetic analyses in mouse fibroblast and melanoma cells demonstrate novel roles for PDGF-AB ligand and PDGF receptor alpha. Scientific Reports, 10, 19303.
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6

Cytoskeletal Protein Immunostaining Assay

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β-Actin (MCA5775GA, AbD Serotec); γ-actin (MCA5776GA, AbD Serotec); p-p44/42 (T202/Y204) (4370S, Cell Signaling); ERK1/2 (4695S, Cell Signaling); p34-Arc (07-227, EMD Millipore); WAVE2 (3659P, Cell Signaling); Cofilin (D3F9) XP®, cofilin1 (5175P, Cell Signaling); p-Cofilin1 (Ser3) (3313, Cell Signaling); N-ras (sc-519, Santa Cruz); PP1α (S3010, Epitomics); Pan-actin (4968, Cell Signaling); α-tubulin (2144, Cell Signaling).
The following secondary Abs were used: FITC-, TRITC-, AlexaFluor488-, AlexaFluor594-, AlexaFluor647-, Cy5-conjugated goat anti-mouse IgG1, IgG2b, IgG and goat anti-rabbit IgG (Southern Biotechnology, Associates Inc., Birmingham, AL; Jackson; Life technologies).
Dugina V., Khromova N., Rybko V., Blizniukov O., Shagieva G., Chaponnier C., Kopnin B, & Kopnin P. (2015). Tumor promotion by γ and suppression by β non-muscle actin isoforms. Oncotarget, 6(16), 14556-14571.
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7

Comprehensive Immunofluorescence Staining Protocol

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Lpd pab 3917,17 (link) Mena mab A351F7D9,54 (link) Wave2 (Cell Signaling Technology), p34Arc (Millipore, 07-227), Tubulin (DM1A), Hsc70 mab (Santa Cruz), GFP mab (Roche), Myc mab (Sigma, 9E10), pTyr mab (Millipore, 4G10) and Vimentin (550513, Biosciences). Alexa-conjugated secondary antibodies, phalloidin (Invitrogen, Biotium) diluted 1:50–1000.
Carmona G., Perera U., Gillett C., Naba A., Law A.L., Sharma V.P., Wang J., Wyckoff J., Balsamo M., Mosis F., De Piano M., Monypenny J., Woodman N., McConnell R.E., Mouneimne G., Van Hemelrijck M., Cao Y., Condeelis J., Hynes R.O., Gertler F.B, & Krause M. (2016). Lamellipodin promotes invasive 3D cancer cell migration via regulated interactions with Ena/VASP and SCAR/WAVE. Oncogene, 35(39), 5155-5169.
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8

Osteoclast Protein Expression Analysis

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After stimulation, whole cell lysates of osteoclasts were lysed in NP-40 buffer containing (150 mM sodium chloride, 1% NP-40, 50 mM Tris pH8) including protease and phosphatase inhibitor cocktail (#11,873,580,001, #P5726, #P004; Sigma Aldrich). Specific antibodies against the following proteins were detected by immunoblotting: cathepsin K (abcam, ab 19,027, 1:1000), integrin alpha-v (abcam, ab179475,1:1000), integrin beta-3 (Cell Signaling #4702,1:1000), DC-STAMP (sc-98769, 1:200), NFATc1 (sc-7294; both from Santa Cruz), Hem1 (Novus Bio, NBP2-13,643, 1:1000), WASP (Santa Cruz, sc-13139, 1:1000) and WAVE2 (Cell Signaling, #3659, 1:1000). For loading control purposes, membranes were stripped and re-probed for GAPDH (Cell Signaling, #3683, 1:2000).
Werbenko E., de Gorter D.J., Kleimann S., Beckmann D., Waltereit-Kracke V., Reinhardt J., Geers F., Paruzel P., Hansen U., Pap T., Stradal T.E, & Dankbar B. (2024). Hem1 is essential for ruffled border formation in osteoclasts and efficient bone resorption. Scientific Reports, 14, 8109.
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9

Modulation of T Cell Actin Dynamics

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Activated T cells were stimulated in the presence of ethanol or GML with plate-bound anti-CD3 on glass coverslips for 15 min. In experiments with inhibitors of cytoskeletal processes, 100 μM Colchine (Cayman Chemicals), 10 μM SMIFH2 (Calbiochem), or 10 μM CK-666 (Calbiochem) solubilized in DMSO were added to the activated T cells at the same time as the GML or solvent. Cells were fixed with 4% paraformaldehyde for 30 min, permeabilized with 0.25% Triton-X for 5 min, and then stained with the appropriate reagents. To detect actin, TMR-Phalloidin (Sigma) was incubated with the fixed and permeabilized cells for 2 hours at 37°C. To detect microcluster formation of signaling proteins, cells were stained with antibodies specific for Arp2 (Cell Signaling), WAVE2 (Cell Signaling), phosphorylated WASp Y290 (Invitrogen), ARPC3 (Millipore), phosphorylated SLP-76 Y128 (BD Pharmingen), and phosphorylated LAT Y191 (Millipore). Staining was detected with the conjugated secondary antibodies DyLight 488 Goat anti-rabbit IgG or Alexa Fluor 568 goat anti-mouse IgG1 (Thermo Fisher). TIRF images were captured by the Leica AM TIRF MC system using a 100x magnification oil immersion objective lens at the University of Iowa Central Microscopy Research Facility. Actin ring structures were imaged using the same microscope using the epifluorescence channels.
Zhang M.S., Tran P.M., Wolff A.J., Tremblay M.M., Fosdick M.G, & Houtman J.C. (2018). Glycerol Monolaurate (GML) induces filopodia formation by disrupting the association between LAT and SLP-76 microclusters. Science signaling, 11(528), eaam9095.
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10

Immunofluorescence Imaging of Cellular Proteins

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Lpd pab 391717 (link), Mena mab A351F7D954 (link), Wave2 (Cell Signaling Technology), p34Arc (Millipore, 07-227), Tubulin (DM1A), Hsc70 mab (Santa Cruz), GFP mab (Roche), Myc mab (Sigma, 9E10), pTyr mab (Millipore, 4G10), Vimentin (550513, Biosciences). Alexa-conjugated secondary antibodies, phalloidin (Invitrogen, Biotium) diluted 1:50-1000.
Carmona G., Perera U., Gillett C., Naba A., Law A.L., Sharma V.P., Wang J., Wyckoff J., Balsamo M., Mosis F., De Piano M., Monypenny J., Woodman N., McConnell R.E., Mouneimne G., Van Hemelrijck M., Cao Y., Condeelis J., Hynes R.O., Gertler F.B, & Krause M. (2016). Lamellipodin promotes invasive 3D cancer cell migration via regulated interactions with Ena/VASP and SCAR/WAVE. Oncogene, 35(39), 5155-5169.
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