Human/Mouse WASF3/WAVE3 Antibody

Detection of Human WASF3/WAVE3 by Western Blot.

Western blot shows lysates of A172 human glioblastoma cell line, SH-SY5Y human neuroblastoma cell line, and LNCaP human prostate cancer cell line. PVDF membrane was probed with 1 µg/mL of Human/Mouse WASF3/WAVE3 Antigen Affinity-purified Polyclonal Antibody (Catalog # AF5515) followed by HRP-conjugated Anti-Goat IgG Secondary Antibody (Catalog # HAF109). A specific band was detected for WASF3/WAVE3 at approximately 70 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.

Detection of Human WASF3/WAVE3 by Western Blot

PPP2R1A interacts with the WAVE Shell Complex (WSC) that contains NHSL1 instead of WAVE proteins.a MCF10A or MDA-MB-231 cells stably expressing FLAG-GFP or FLAG-GFP PPP2R1A were subjected to FLAG-GFP Tandem Affinity Purification and purified proteins were resolved by SDS–PAGE and silver stained. The same samples were used for mass spectrometry and Western blots. Three biological repeats with similar results. b Label-free quantification (LFQ) of proteins identified by mass spectrometry. In both cell lines, NHSL1 is present at the expense of WAVE subunits. Log10 of LFQ values, mean ± sem of three biological repeats. c GFP immunoprecipitates were analyzed by Western blots with the indicated antibodies. Three biological repeats with similar results. d PPP2R1A associates with the WSC through NHSL1. MCF10A cells expressing FLAG-GFP PPP2R1A were transfected with pools of siRNAs targeting NHSL1 or non-targeting siRNAs (CTRL). GFP immunoprecipitates were analyzed by western blots. Three biological repeats with similar results. e A MCF10A cell line stably transfected with two plasmids expressing FLAG-GFP PPP2R1A and PC-HA BRK1 was subjected to FLAG-PC Tandem Affinity Purification to purify the WSC. WSC composition was analyzed by mass spectrometry. Silver stained SDS–PAGE of purified WSC. The identity of WSC subunits and the lack of WAVE2 was confirmed using Western blots. 3 biological repeats with similar results. f Comparison of the structural model of the WSC obtained using AlphaFold2 with the X-ray crystal structure WAVE Regulatory Complex (PDB:3P8C [https://www.wwpdb.org/pdb?id=pdb_00003p8c]). The WSC was composed of NHSL1 (1-123) in purple, CYFIP1 in green, NCKAP1 in cyan, BRK1 in yellow and ABI2 (1–160) in salmon. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/37322026), licensed under a CC-BY license. Not internally tested by R&D Systems.

Detection of Human WASF3/WAVE3 by Western Blot

PPP2R1A interacts with the WAVE Shell Complex (WSC) that contains NHSL1 instead of WAVE proteins.a MCF10A or MDA-MB-231 cells stably expressing FLAG-GFP or FLAG-GFP PPP2R1A were subjected to FLAG-GFP Tandem Affinity Purification and purified proteins were resolved by SDS–PAGE and silver stained. The same samples were used for mass spectrometry and Western blots. Three biological repeats with similar results. b Label-free quantification (LFQ) of proteins identified by mass spectrometry. In both cell lines, NHSL1 is present at the expense of WAVE subunits. Log10 of LFQ values, mean ± sem of three biological repeats. c GFP immunoprecipitates were analyzed by Western blots with the indicated antibodies. Three biological repeats with similar results. d PPP2R1A associates with the WSC through NHSL1. MCF10A cells expressing FLAG-GFP PPP2R1A were transfected with pools of siRNAs targeting NHSL1 or non-targeting siRNAs (CTRL). GFP immunoprecipitates were analyzed by western blots. Three biological repeats with similar results. e A MCF10A cell line stably transfected with two plasmids expressing FLAG-GFP PPP2R1A and PC-HA BRK1 was subjected to FLAG-PC Tandem Affinity Purification to purify the WSC. WSC composition was analyzed by mass spectrometry. Silver stained SDS–PAGE of purified WSC. The identity of WSC subunits and the lack of WAVE2 was confirmed using Western blots. 3 biological repeats with similar results. f Comparison of the structural model of the WSC obtained using AlphaFold2 with the X-ray crystal structure WAVE Regulatory Complex (PDB:3P8C [https://www.wwpdb.org/pdb?id=pdb_00003p8c]). The WSC was composed of NHSL1 (1-123) in purple, CYFIP1 in green, NCKAP1 in cyan, BRK1 in yellow and ABI2 (1–160) in salmon. Image collected and cropped by CiteAb from the following open publication (https://pubmed.ncbi.nlm.nih.gov/37322026), licensed under a CC-BY license. Not internally tested by R&D Systems.