vimentin antibody  (NSJ Bioreagents)

Journal: Non-coding RNA Research

Article Title: SNHG5 enhances colorectal cancer metastasis through RNA–protein interaction with GNB2 and activation of canonical Wnt signaling

doi: 10.1016/j.ncrna.2025.12.002

Figure Lengend Snippet: Highly metastatic MC38-derived CRC cells exhibit enhanced proliferative, migratory, and invasive properties with mesenchymal characteristics . (A) CCK-8 assay demonstrated significantly higher proliferative capacity in the highly metastatic MC38-F3 subline compared to the parental low-metastatic MC38-F0 cells over a 96-h time course. (B) Representative colony formation images and quantification revealed increased clonogenicity in the MC38-F3 subline. (C) Wound healing assays indicated enhanced migratory ability in MC38-F3 cells at 48 h post-scratch. (D) Transwell migration and Matrigel-coated invasion assays showed that MC38-F3 cells exhibited significantly increased motility and invasiveness. Scale bars = 100 μm. (E) Western blot analysis revealed downregulation of the epithelial marker E-cadherin and upregulation of mesenchymal markers N-cadherin, Vimentin, and Slug in MC38-F3 cells. (F) RT-qPCR analysis confirmed significant upregulation of EMT-associated transcription factors (Snail, Slug, Twist1, ZEB1, ZEB2) in MC38-F3 cells. (G) Immunofluorescence staining corroborated the EMT phenotype, showing reduced E-cadherin and increased Vimentin expression in MC38-F3 cells. Nuclei were counterstained with DAPI. Scale bars = 20 μm. Data are presented as mean ± SEM from at least three independent experiments. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001 (Student's t -test).

Article Snippet: Membranes were blocked in 5 % non-fat dry milk diluted in TBST (Tris-buffered saline containing 0.1 % Tween-20) for 1 h at room temperature and incubated overnight at 4 °C with the following primary antibodies: GNB2 (1:1000; rabbit monoclonal, clone EP3262Y, Abcam, ab108504), E-cadherin (1:1000; rabbit monoclonal, CST, #14472), N-cadherin (1:1000; rabbit monoclonal, CST, #13116), Vimentin (1:1000; rabbit polyclonal, Proteintech, 10366-1-AP), Slug (1:1000; rabbit monoclonal, CST, #9585) and GAPDH (1:5000; mouse monoclonal, Proteintech, 60004-1-Ig) as a loading control.

Techniques: Derivative Assay, CCK-8 Assay, Migration, Western Blot, Marker, Quantitative RT-PCR, Immunofluorescence, Staining, Expressing

Journal: Non-coding RNA Research

Article Title: SNHG5 enhances colorectal cancer metastasis through RNA–protein interaction with GNB2 and activation of canonical Wnt signaling

doi: 10.1016/j.ncrna.2025.12.002

Figure Lengend Snippet: The Snhg5–GNB2 axis promotes EMT and activates Wnt/β-catenin signaling in CRC cells . (A) Immunohistochemistry of liver metastatic tissues revealed that Snhg5 knockdown reduced β-catenin and Vimentin expression while increasing E-cadherin levels. These changes were partially reversed by GNB2 overexpression. Quantification of IHC signal intensity (IOD) supported the observed protein alterations. Scale bars = 100 μm. (B, D) Western blot analysis of MC38-F0 (B) and MC38-F3 (D) cells showed that Snhg5 silencing increased E-cadherin expression while decreasing N-cadherin, Vimentin, and Slug levels. GNB2 overexpression reversed these EMT-associated changes. (C, E) RT-qPCR confirmed that knockdown of Snhg5 suppressed the transcription of key EMT regulators (Snail, Slug, Twist1, ZEB1, ZEB2), which was restored by GNB2 overexpression in both cell models. (F, H) Western blot analysis of Wnt signaling components revealed that Snhg5 knockdown reduced total β-catenin and phosphorylated GSK-3β (Ser9) levels, with no significant change in total GSK-3β. GNB2 overexpression reversed these effects in both MC38-F0 and F3 cells. (G, I) RT-qPCR analysis demonstrated that Snhg5 depletion led to reduced expression of canonical Wnt target genes (AXIN2, c-MYC, Cyclin D1), which was significantly restored by GNB2. Data are presented as mean ± SEM from at least three independent experiments. ∗ P < 0.05, ∗∗ P < 0.01, ∗∗∗ P < 0.001 (one-way ANOVA with Tukey's post hoc test).

Article Snippet: Membranes were blocked in 5 % non-fat dry milk diluted in TBST (Tris-buffered saline containing 0.1 % Tween-20) for 1 h at room temperature and incubated overnight at 4 °C with the following primary antibodies: GNB2 (1:1000; rabbit monoclonal, clone EP3262Y, Abcam, ab108504), E-cadherin (1:1000; rabbit monoclonal, CST, #14472), N-cadherin (1:1000; rabbit monoclonal, CST, #13116), Vimentin (1:1000; rabbit polyclonal, Proteintech, 10366-1-AP), Slug (1:1000; rabbit monoclonal, CST, #9585) and GAPDH (1:5000; mouse monoclonal, Proteintech, 60004-1-Ig) as a loading control.

Techniques: Immunohistochemistry, Knockdown, Expressing, Over Expression, Western Blot, Quantitative RT-PCR