nucleolin polyclonal antibody  (Proteintech)

Journal: Biomolecules

Article Title: Myogenic Anti-Nucleolin Aptamer iSN04 Inhibits Proliferation and Promotes Differentiation of Vascular Smooth Muscle Cells

doi: 10.3390/biom14060709

Figure Lengend Snippet: Nucleolin localization and iSN04 incorporation in A10 cells. ( A ) Representative fluorescence images of nucleolin staining of A10 cells in GM (day 0) and DM with or without 10 μM iSN04 (day 4). Scale bar, 50 μm. ( B ) Representative fluorescence images of A10 cells treated with 5 μg/mL 6-FAM-iSN04 in GM. Scale bar, 50 μm. 6-FAM, 6-carboxyfluorescein; DAPI, 4′,6-diamidino-2-phenylindole; DM, differentiation medium; GM, growth medium.

Article Snippet: The cells were fixed with 2% paraformaldehyde, permeabilized with 0.2% Triton X-100, and immunostained with 1.0 μg/mL rabbit polyclonal anti-nucleolin antibody (ab22758; Abcam, Cambridge, UK) or 1.0 μg/mL mouse monoclonal anti-α-SMA antibody (1A4, ab7817; Abcam) overnight at 4 °C.

Techniques: Fluorescence, Staining

Journal: Biomolecules

Article Title: Myogenic Anti-Nucleolin Aptamer iSN04 Inhibits Proliferation and Promotes Differentiation of Vascular Smooth Muscle Cells

doi: 10.3390/biom14060709

Figure Lengend Snippet: The effect of iSN04 on proliferation and differentiation of hAoSMCs. ( A ) Representative fluorescence images of nucleolin staining of hAoSMCs in hGM (day 0) and hDM (day 4). Scale bar, 50 μm. ( B ) Representative fluorescence images of EdU staining of hAoSMCs pre-treated with 30 μM iSN04 in hGM for 48 h and then with 10 μM EdU in hGM for 12 h. Scale bar, 200 μm. The ratio of EdU + cells were quantified. ** p < 0.01 vs. control (Student’s t -test). n = 4. ( C ) qPCR results of a cell-cycle marker gene, Ki-67 ( MKI67 ), and contractile SMC marker genes, α-SMA ( ACTA2 ), SM22α ( TAGLN ), and caldesmon ( CALD1 ), in hAoSMCs treated with 30 μM iSN04 in hGM for 48 h. * p < 0.05; ** p < 0.01 vs. control (Student’s t -test). n = 3. DAPI, 4′,6-diamidino-2-phenylindole; DM, differentiation medium; EdU, 5-ethynyl-2′-deoxyuridine; GM, growth medium; hAoSMC, human aortic smooth muscle cell; hDM, differentiation medium for human aortic smooth muscle cell; hGM, growth medium for human aortic smooth muscle cell; qPCR, quantitative real-time RT-PCR; SMC, smooth muscle cell.

Article Snippet: The cells were fixed with 2% paraformaldehyde, permeabilized with 0.2% Triton X-100, and immunostained with 1.0 μg/mL rabbit polyclonal anti-nucleolin antibody (ab22758; Abcam, Cambridge, UK) or 1.0 μg/mL mouse monoclonal anti-α-SMA antibody (1A4, ab7817; Abcam) overnight at 4 °C.

Techniques: Fluorescence, Staining, Control, Marker, Quantitative RT-PCR

Journal: bioRxiv

Article Title: Impaired migration and metastatic spread of human melanoma by a novel small molecule targeting the transmembrane domain of death receptor p75 NTR

doi: 10.1101/2023.11.13.566904

Figure Lengend Snippet: (A) Schematic shows the experimental design for the assessment of a human melanoma xenograft lung metastasis model following treatment with Np75-4A22. (B) Live-monitoring of lung metastatic tumor growth of A875-NT melanoma cells using the Xenogen IVIS-200 luciferase imaging system. Bioluminescent imaging of luciferase expressing A875-NT melanoma cells (control) in the lungs of SCID mice was measured 1, 2 and 3-weeks following treatment with Np75-4A22 (200 mg/kg/day) or vehicle alone. Representative images taken 3 weeks following implantation are shown (left panel). Graph shows the average quantification of luciferase signal (N=8-10 mice/group; right panel). Shown is one representative experiment from N=3 independent experiments. Data are expressed as the mean ± SEM. Means that are significantly different between treatments as analyzed by Student t-test are indicated. *, p<0.05: **, p<0.01. (C) Histological analysis of lung tumor burden in SCID mice bearing A875-NT or A875-shp75 tumors 3 weeks following of treatment with Np75-4A22 or vehicle alone. Left panel shows representative images of formalin fixed paraffin embedded lung sections stained by immunohistochemistry with a human-specific nucleolin antibody (brown) to detect the presence of tumor cells. Sections were counterstained with hematoxylin (blue). Scale bar, 60μm. Right panel shows quantification of the number of cancer cells per lung tissue section. Five non-sequential lung sections, 100 μm apart, were counted and averaged for each mouse (N=8-10 mice per group). Data is expressed as the mean ± SEM. Means that are significantly different are indicated. A875-NT vehicle vs Np75-4A22 ** p=0.0179; A875-NT vs A875-shp75 # p=0.0367; A857-shp75 vehicle vs Np75-4A22 not significant (n.s.), one-way ANOVA with Tukey’s multiple comparisons test.

Article Snippet: Human-specific mouse polyclonal nucleolin antibody was from Abcam (#ab136649).

Techniques: Luciferase, Imaging, Expressing, Formalin-fixed Paraffin-Embedded, Staining, Immunohistochemistry

Journal: bioRxiv

Article Title: Impaired migration and metastatic spread of human melanoma by a novel small molecule targeting the transmembrane domain of death receptor p75 NTR

doi: 10.1101/2023.11.13.566904

Figure Lengend Snippet: p75 NTR immunostaining (red) in lung metastasis induced by A875-NT (A) or shp75-A875 (B) cells counter-stained for human nucleolin (green) and DAPI (white). Scale bar, 100μM.

Article Snippet: Human-specific mouse polyclonal nucleolin antibody was from Abcam (#ab136649).

Techniques: Immunostaining, Staining

Journal: PLoS ONE

Article Title: Proteomic Analysis of Chikungunya Virus Infected Microgial Cells

doi: 10.1371/journal.pone.0034800

Figure Lengend Snippet: CHME-5 cells either mock infected or infected with CHIKV at MOI 0.1 were collected at day 2 p.i. (A, B, D) or on days 1 to 3 p.i. (C) and subsequently (A) stained with an anti-alphavirus antibody and the percentage of infected cells analyzed by flow cytometry or (B) stained with Annexin V-FITC and PI and the percentage of apoptotic cells analyzed by flow cytometery or (C, D) used for total protein extraction and (C) analyzed by western blotting with an anti-alphavirus monoclonal antibody and an anti-actin polyclonal antibody or (D) the differential proteome determined by 2D-PAGE. Representative gels from 6 biological replicates are shown. (A and B) Bar graphs represent the means ± SD of 6 replications.

Article Snippet: Antibodies used included a 1∶400 dilution of a rabbit anti-JAK 2 polyclonal antibody (sc-278; Santa Cruz Biotechnology Inc.), a 1∶6,000 dilution of a rabbit anti-Hsp90 polyclonal antibody (sc-7947; Santa Cruz Biotechnology Inc.), a 1∶4,000 dilution of a rabbit anti-Hsp70 polyclonal antibody (sc-1060; Santa Cruz Biotechnology Inc.), a 1∶5,000 dilution of a rabbit anti-nucleolin polyclonal antibody (sc-13057; Santa Cruz Biotechnology Inc.), a 1∶3,000 dilution of a rabbit anti-heterogeneous nuclear ribonucleoprotein polyclonal antibody (ab65049; Abcam, Cambridge, MA), a 1∶10,000 dilution of a rabbit anti-elongation factor 2 polyclonal antibody (ab33523; Abcam) all followed by a 1∶4,000 dilution of a goat horseradish peroxidase (HRP) conjugated anti-rabbit IgG polyclonal antibody (31460; Pierce, Rockford, IL) as well as a 1∶5,000 dilution of a goat anti-actin polyclonal antibody (sc-1616; Santa Cruz Biotechnology Inc.) followed by a 1∶8,000 dilution of a rabbit HRP conjugated anti-goat IgG polyclonal antibody (31402; Pierce), as well as a 1∶1000 dilution of a mouse anti-alphavirus monoclonal antibody (Santa Cruz Biotechnology Inc., Santa Cruz, CA) followed by a 1∶5,000 dilution of a rabbit HRP conjugated anti-mouse IgG polyclonal antibody (A9044; Sigma, Sigma-Aldrich, St Louis, MO).

Techniques: Infection, Staining, Flow Cytometry, Protein Extraction, Western Blot

Journal: PLoS ONE

Article Title: Proteomic Analysis of Chikungunya Virus Infected Microgial Cells

doi: 10.1371/journal.pone.0034800

Figure Lengend Snippet: (A) CHME-5 cells were either mock infected or infected with CHIKV at MOI 0.1 before extraction of proteins and analysis by Western blot analysis on 1 and 2 d.p.i. hnRNP: heterogeneous nuclear ribonucleoprotein; NCL: nucleolin; JAK2: tyrosine-protein kinase JAK2; Hsp70: heat shock protein 70; Hsp90: heat shock protein 90. (B). CHME-5 cells were either mock infected or infected with CHIKV at MOI 0.1 before extraction of total RNA and analysis by RT-PCR on 1, 2 and 3 d.p.i. BRE1B: E3 ubiquitin-protein ligase; CUL9: Cullin-9; CHD2: chromodomain-helicase-DNA binding protein 2; MTERF: mitochondrial precursor transcription termination factor; ROD1: regulator of differentiation 1 isoform; PIK3CD: phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit delta; GCDH: mitochondrial glutaryl-CoA dehydrogenase isoform precursor; HSDL2: hydroxysteroid dehydrogenase-like protein 2; PLCH2: 1-phosphatidylinositol-4,5-bisphosphate phosphodiesterase eta-2; ALOX12: 12-lipoxygenase; DRPLA: Dentatorubral pallidoluysian atrophy protein; DENND3: DENN domain-containing protein 3; HIS1H2B: Histone 2B.

Article Snippet: Antibodies used included a 1∶400 dilution of a rabbit anti-JAK 2 polyclonal antibody (sc-278; Santa Cruz Biotechnology Inc.), a 1∶6,000 dilution of a rabbit anti-Hsp90 polyclonal antibody (sc-7947; Santa Cruz Biotechnology Inc.), a 1∶4,000 dilution of a rabbit anti-Hsp70 polyclonal antibody (sc-1060; Santa Cruz Biotechnology Inc.), a 1∶5,000 dilution of a rabbit anti-nucleolin polyclonal antibody (sc-13057; Santa Cruz Biotechnology Inc.), a 1∶3,000 dilution of a rabbit anti-heterogeneous nuclear ribonucleoprotein polyclonal antibody (ab65049; Abcam, Cambridge, MA), a 1∶10,000 dilution of a rabbit anti-elongation factor 2 polyclonal antibody (ab33523; Abcam) all followed by a 1∶4,000 dilution of a goat horseradish peroxidase (HRP) conjugated anti-rabbit IgG polyclonal antibody (31460; Pierce, Rockford, IL) as well as a 1∶5,000 dilution of a goat anti-actin polyclonal antibody (sc-1616; Santa Cruz Biotechnology Inc.) followed by a 1∶8,000 dilution of a rabbit HRP conjugated anti-goat IgG polyclonal antibody (31402; Pierce), as well as a 1∶1000 dilution of a mouse anti-alphavirus monoclonal antibody (Santa Cruz Biotechnology Inc., Santa Cruz, CA) followed by a 1∶5,000 dilution of a rabbit HRP conjugated anti-mouse IgG polyclonal antibody (A9044; Sigma, Sigma-Aldrich, St Louis, MO).

Techniques: Infection, Western Blot, Reverse Transcription Polymerase Chain Reaction, Binding Assay

Journal: PLoS ONE

Article Title: Proteomic Analysis of Chikungunya Virus Infected Microgial Cells

doi: 10.1371/journal.pone.0034800

Figure Lengend Snippet: Values of fold change in response to CHIKV infection, significance and half life of selected proteins.

Article Snippet: Antibodies used included a 1∶400 dilution of a rabbit anti-JAK 2 polyclonal antibody (sc-278; Santa Cruz Biotechnology Inc.), a 1∶6,000 dilution of a rabbit anti-Hsp90 polyclonal antibody (sc-7947; Santa Cruz Biotechnology Inc.), a 1∶4,000 dilution of a rabbit anti-Hsp70 polyclonal antibody (sc-1060; Santa Cruz Biotechnology Inc.), a 1∶5,000 dilution of a rabbit anti-nucleolin polyclonal antibody (sc-13057; Santa Cruz Biotechnology Inc.), a 1∶3,000 dilution of a rabbit anti-heterogeneous nuclear ribonucleoprotein polyclonal antibody (ab65049; Abcam, Cambridge, MA), a 1∶10,000 dilution of a rabbit anti-elongation factor 2 polyclonal antibody (ab33523; Abcam) all followed by a 1∶4,000 dilution of a goat horseradish peroxidase (HRP) conjugated anti-rabbit IgG polyclonal antibody (31460; Pierce, Rockford, IL) as well as a 1∶5,000 dilution of a goat anti-actin polyclonal antibody (sc-1616; Santa Cruz Biotechnology Inc.) followed by a 1∶8,000 dilution of a rabbit HRP conjugated anti-goat IgG polyclonal antibody (31402; Pierce), as well as a 1∶1000 dilution of a mouse anti-alphavirus monoclonal antibody (Santa Cruz Biotechnology Inc., Santa Cruz, CA) followed by a 1∶5,000 dilution of a rabbit HRP conjugated anti-mouse IgG polyclonal antibody (A9044; Sigma, Sigma-Aldrich, St Louis, MO).

Techniques: Infection