antibodies against e2f2  (Novus Biologicals)

Journal: Experimental and Therapeutic Medicine

Article Title: LncRNA ZEB1-AS1 reduces liver cancer cell proliferation by targeting miR-365a-3p

doi: 10.3892/etm.2019.7358

Figure Lengend Snippet: Regulation of E2F2 by miR-365a-3p. (A) Schematic diagram presenting the putative miR-365a-3p response element on the E2F2 3′UTR. The miR-365a-3p seed sequence region is marked in pink. (B) Schematic diagram of the luciferase reporter vector constructs utilized. (C) Luciferase activity in 293T cells co-transfected miRNA mimics, empty luciferase reporter pmirGLOs and luciferase reporter pmirGLOs containing E2F2 3′ UTR WT or E2F2 3′UTR Mut. ***P<0.001 vs. WT+miR-NC. (D) RT-qPCR analysis of miR-365a-3p levels in HepG2 and HCCLM6 cells transfected with miR-365a-3p mimics or miR-NC. **P<0.01 and ***P<0.001 vs. miR-NC. (E) RT-qPCR analysis of E2F2 mRNA levels in HepG2 and HCCLM6 cells transfected with miR-365a-3p mimics or miR-NC. **P<0.01 and ***P<0.001 vs. miR-NC. (F) Western blot analysis of E2F2 in HepG2 and HCCLM6 cells transfected with miR-365a-3p mimics or miR-NC. Data are presented as the mean ± standard deviation (n=3 miR or miRNA, microRNA; UTR, untranslated region; WT, wild type; mut, mutant; NC, negative control; RT-qPCR, reverse transcription quantitative polymerase chain reaction.

Article Snippet: The membranes were incubated with primary antibodies against E2F2 (1:500; cat. no. SAB4500684; Sigma-Aldrich; Merck KGaA) and GAPDH (1:1,000; cat. no. 2118; Cell Signaling Technology, Inc., Danvers, MA, USA) overnight at 4°C.

Techniques: Sequencing, Luciferase, Plasmid Preparation, Construct, Activity Assay, Transfection, Quantitative RT-PCR, Western Blot, Standard Deviation, Mutagenesis, Negative Control, Real-time Polymerase Chain Reaction

Journal: Experimental and Therapeutic Medicine

Article Title: LncRNA ZEB1-AS1 reduces liver cancer cell proliferation by targeting miR-365a-3p

doi: 10.3892/etm.2019.7358

Figure Lengend Snippet: Regulation of E2F2 by ZEB1-AS1. (A) RT-qPCR analysis of E2F2 mRNA levels in HepG2 and HCCLM6 cells transfected with indicated plasmids. (B) E2F2 expression in HepG2 and HCCLM6 cells was determined via western blotting (C) RT-qPCR analysis of E2F2 mRNA levels in 32 pairs of hepatocarcinoma tissue and matched adjacent normal tissue samples. (D) Pearson's correlation analysis of ZEB1-AS1 and E2F2 mRNA levels in 32 hepatocarcinoma tissue samples. Data are expressed as the mean ± standard deviation (n=3). **P<0.01 vs. sh-NC. ZEB1-AS1, zinc-finger E-box binding homeobox 1 antisense 1; RT-qPCR, reverse transcription-quantitative polymerase chain reaction; sh, short hairpin; NC, negative control; HCC, hepatocarcinoma.

Article Snippet: The membranes were incubated with primary antibodies against E2F2 (1:500; cat. no. SAB4500684; Sigma-Aldrich; Merck KGaA) and GAPDH (1:1,000; cat. no. 2118; Cell Signaling Technology, Inc., Danvers, MA, USA) overnight at 4°C.

Techniques: Quantitative RT-PCR, Transfection, Expressing, Western Blot, Standard Deviation, Binding Assay, Real-time Polymerase Chain Reaction, Negative Control

Journal: Experimental and Therapeutic Medicine

Article Title: LncRNA ZEB1-AS1 reduces liver cancer cell proliferation by targeting miR-365a-3p

doi: 10.3892/etm.2019.7358

Figure Lengend Snippet: Regulation of E2F2 expression and cell proliferation via ZEB1-AS1 and miR-365a-3p. (A) RT-qPCR analysis of miR-365a-3p levels in HepG2 cells co-transfected with indicated plasmids and miRNA inhibitors. (B) RT-qPCR analysis of E2F2 mRNA levels in HepG2 cells co-transfected with indicated plasmids and miRNA inhibitors. (C) Western blot analysis of E2F2 expression in HepG2 cells co-transfected with indicated plasmids and miRNA inhibitors. (D) Cell Counting Kit-8 analysis of HepG2 cell viabilities presented as a cell proliferation curve. (E) Flow cytometric analysis of HepG2 cell cycle distribution. All data are presented as the mean ± standard deviation. *P<0.05 and **P<0.01. ZEB1-AS1, zinc-finger E-box binding homeobox 1 antisense 1; miR and miRNA, microRNA; inh, inhibitor; NC, negative control; sh, short hairpin.

Article Snippet: The membranes were incubated with primary antibodies against E2F2 (1:500; cat. no. SAB4500684; Sigma-Aldrich; Merck KGaA) and GAPDH (1:1,000; cat. no. 2118; Cell Signaling Technology, Inc., Danvers, MA, USA) overnight at 4°C.

Techniques: Expressing, Quantitative RT-PCR, Transfection, Western Blot, Cell Counting, Standard Deviation, Binding Assay, Negative Control

Journal: PPAR Research

Article Title: PPAR- γ Ligand Inhibits Nasopharyngeal Carcinoma Cell Proliferation and Metastasis by Regulating E2F2

doi: 10.1155/2019/8679271

Figure Lengend Snippet: Expressions of E2F2 and PPAR- γ in clinical samples of undifferentiated NPC and NPG and associations with clinicopathological characteristics of undifferentiated NPC samples.

Article Snippet: After three washes with phosphate-buffered saline (PBS), the sections were incubated in a blocking solution containing bovine serum albumin (BSA) for 20 min, followed by exposure to primary antibodies against E2F2 (YT1443, Immunoway, rabbit polyclonal, 1:200 dilution) and PPAR- γ (Novusbio, rabbit polyclonal, 1:100 dilution) at 4°C overnight.

Techniques: Expressing

Journal: PPAR Research

Article Title: PPAR- γ Ligand Inhibits Nasopharyngeal Carcinoma Cell Proliferation and Metastasis by Regulating E2F2

doi: 10.1155/2019/8679271

Figure Lengend Snippet: E2F2 and PPAR-γ protein expression in nonkeratinizing nasopharyngeal carcinoma (NPC) and nasopharyngitis (NPG) tissues . Immunohistochemistry was performed to detect E2F2 and PPAR- γ protein expression in nonkeratinizing NPC tissues (a, b) and NPG tissues (c, d). Scale bar, 50 μ m (e, f). The immunohistochemistry data were analyzed semiquantitatively to determine the E2F2 and PPAR- γ expression levels in nonkeratinizing NPC and NPG tissues. Data are shown as means ± standard deviations. ∗∗ P <0.01.

Article Snippet: After three washes with phosphate-buffered saline (PBS), the sections were incubated in a blocking solution containing bovine serum albumin (BSA) for 20 min, followed by exposure to primary antibodies against E2F2 (YT1443, Immunoway, rabbit polyclonal, 1:200 dilution) and PPAR- γ (Novusbio, rabbit polyclonal, 1:100 dilution) at 4°C overnight.

Techniques: Expressing, Immunohistochemistry

Journal: PPAR Research

Article Title: PPAR- γ Ligand Inhibits Nasopharyngeal Carcinoma Cell Proliferation and Metastasis by Regulating E2F2

doi: 10.1155/2019/8679271

Figure Lengend Snippet: E2F2 and PPAR-γ protein expression in nonkeratinizing nasopharyngeal carcinoma (NPC) and nasopharyngitis (NPG) tissue lysates . (a, b) RT-PCR and Western blotting were performed to detect the expression of E2F2, PPAR- γ , and GAPDH in lysates of nonkeratinizing NPC and NPG tissues. (c, d) Quantitative analyses of the E2F2 and PPAR- γ expression levels in nonkeratinizing NPC and NPG tissues. Data are expressed as means ± standard deviations. ∗ P <0.05; ∗∗ P <0.01.

Article Snippet: After three washes with phosphate-buffered saline (PBS), the sections were incubated in a blocking solution containing bovine serum albumin (BSA) for 20 min, followed by exposure to primary antibodies against E2F2 (YT1443, Immunoway, rabbit polyclonal, 1:200 dilution) and PPAR- γ (Novusbio, rabbit polyclonal, 1:100 dilution) at 4°C overnight.

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Western Blot

Journal: PPAR Research

Article Title: PPAR- γ Ligand Inhibits Nasopharyngeal Carcinoma Cell Proliferation and Metastasis by Regulating E2F2

doi: 10.1155/2019/8679271

Figure Lengend Snippet: E2F2 and PPAR-γ expression in CNE1 and CNE2 nasopharyngeal carcinoma cells after treatment with PPAR-γ ligand . (a, b) Western blotting was performed to detect the expression of E2F2, PPAR- γ , and GAPDH in CNE1 and CNE2 cells after treatment with dose-dependent PPAR- γ agonist Rog. (c, d) Western blotting was performed to detect the expression of E2F2, PPAR- γ , and GAPDH in CNE1 and CNE2 cells after treatment with PPAR- γ agonist (rosiglitazone, Rog) and PPAR- γ antagonist (GW9662).

Article Snippet: After three washes with phosphate-buffered saline (PBS), the sections were incubated in a blocking solution containing bovine serum albumin (BSA) for 20 min, followed by exposure to primary antibodies against E2F2 (YT1443, Immunoway, rabbit polyclonal, 1:200 dilution) and PPAR- γ (Novusbio, rabbit polyclonal, 1:100 dilution) at 4°C overnight.

Techniques: Expressing, Western Blot

Journal: Oncotarget

Article Title: miR-155 regulates the proliferation and invasion of clear cell renal cell carcinoma cells by targeting E2F2

doi: 10.18632/oncotarget.7951

Figure Lengend Snippet: ( A ) miR-155 expression levels were significantly upregulated in ccRCC cancer tissues compared with normal tissues. ( B – D ) Correlations between miR-155 expression level and the clinical stage, T stage, Fuhrman grades of ccRCC respectively. ( E ) miR-155 expressions in various RCC cell lines compared with that in HKC. ( F ) Downregulation of E2F2 mRNA levels in ccRCC cancer tissues; here, normal tissues served as controls. ( G ) Correlations between E2F2 expression level and the clinical stage of ccRCC. ( H ) Negative correlation of E2F2 mRNA levels and miR-155 levels ( n = 54, r 2 = 0.4121, P < 0.0001). ( I ) Representative images of E2F2 IHC in ccRCC cancer tissues and their paired normal tissues. ( J ) Western blot of E2F2 showed alterations in protein levels consistent with variations in mRNA levels in clinical samples. Data represent the mean ± SD. (* P < 0.05; ** P < 0.01; *** P < 0.001).

Article Snippet: Membranes were blocked and then incubated with primary antibodies against E2F2 (Santa Cruz, sc-632, USA), N-Cadherin (Abcam, Ab76057), E-Cadherin (Cell Signaling Technology, #3195, USA), Vimentin (Cell Signaling Technology, #5741, USA), ZEB1 (Cell Signaling Technology#3396, USA), or β-actin (ZSGB-BIO, TA-09, China) followed by incubation with horseradish peroxidase-conjugated secondary antibodies.

Techniques: Expressing, Western Blot

Journal: Oncotarget

Article Title: miR-155 regulates the proliferation and invasion of clear cell renal cell carcinoma cells by targeting E2F2

doi: 10.18632/oncotarget.7951

Figure Lengend Snippet: ( A ) Alteration of E2F2 expression levels in 786-O cells 48 h after siRNA treatment compared with siNC. ( B ) Alteration of E2F2 expression levels in ACHN cells 48 h after transfection with lentiviral E2F2 plasmids compared with the empty vector. ( C ) MTS assay revealed that transfection of the siRNA of E2F2 significantly accelerated proliferation velocity in 786-O cells while E2F2 overexpression attenuated proliferation in ACHN cells. ( D ) Effect of E2F2 overexpression or suppression on the colony formation of ACHN and 786-O cells, respectively. The number of foci of > 50 cells was counted after 14 d. ( E ) Representative photographs of transwell assays (magnification, ×100) of 786-O and ACHN cells identifying E2F2 as a tumor suppressor. ( F ) E2F2 knockdown in 786-O cells significantly increased the number of viable cells; in ACHN cells, E2F2 overexpression largely decreased cell viability. Data represent the mean ± SD. Each experiment was performed in triplicate. (* P < 0.05; ** P < 0.01; *** P < 0.001).

Article Snippet: Membranes were blocked and then incubated with primary antibodies against E2F2 (Santa Cruz, sc-632, USA), N-Cadherin (Abcam, Ab76057), E-Cadherin (Cell Signaling Technology, #3195, USA), Vimentin (Cell Signaling Technology, #5741, USA), ZEB1 (Cell Signaling Technology#3396, USA), or β-actin (ZSGB-BIO, TA-09, China) followed by incubation with horseradish peroxidase-conjugated secondary antibodies.

Techniques: Expressing, Transfection, Plasmid Preparation, MTS Assay, Over Expression, Knockdown

Journal: Oncotarget

Article Title: miR-155 regulates the proliferation and invasion of clear cell renal cell carcinoma cells by targeting E2F2

doi: 10.18632/oncotarget.7951

Figure Lengend Snippet: ( A ) Sequence alignment of the E2F2 3′UTR with wild-type (WT) versus mutant (MUT) potential miR-155 targeting sites. ( B ) mRNA levels of E2F2 were examined by qRT-PCR in 786-O and ACHN cells with different interferences. ( C ) Transfection of the miR-155 mimic or miR-155 inhibitor in 786-O and ACHN greatly changed E2F2 protein levels. ( D ) Immunofluorescence staining results showed the inverse effect of miR-155 on E2F2 in different cell lines. ( E ) Luciferase reporter assay showed decreased reporter activity after transfection of the wild-type E2F2 3′UTR reporter construct in 293T cells overexpressing miR-155. The E2F2 3′UTR MUT and control constructs showed no effect on reporter activity. Here, the Renilla luciferase construct was used as an internal control. The normalized luciferase activity of the control construct in each experiment was set to 1. Data represent the mean ± SD. (* P < 0.05; ** P < 0.01; *** P < 0.001).

Article Snippet: Membranes were blocked and then incubated with primary antibodies against E2F2 (Santa Cruz, sc-632, USA), N-Cadherin (Abcam, Ab76057), E-Cadherin (Cell Signaling Technology, #3195, USA), Vimentin (Cell Signaling Technology, #5741, USA), ZEB1 (Cell Signaling Technology#3396, USA), or β-actin (ZSGB-BIO, TA-09, China) followed by incubation with horseradish peroxidase-conjugated secondary antibodies.

Techniques: Sequencing, Mutagenesis, Quantitative RT-PCR, Transfection, Immunofluorescence, Staining, Luciferase, Reporter Assay, Activity Assay, Construct, Control

Journal: Oncotarget

Article Title: miR-155 regulates the proliferation and invasion of clear cell renal cell carcinoma cells by targeting E2F2

doi: 10.18632/oncotarget.7951

Figure Lengend Snippet: ( A ) miR-155 and E2F2 mRNA level changes in 786-O cells with the miR-155 mimic (versus the control) and the E2F2 plasmid (versus the control) 48 h after transfection. ( B ) The mRNA levels of miR-155 and E2F2 in ACHN were analyzed by qRT-PCR after the use of miR-155 inhibitor and siE2F2. ( C ) E2F2 protein level alterations in 786-O cells with the miR-155 mimic (versus the control) and the E2F2 plasmid (versus the control) 48 h after transfection. After co-transfection of ACHN cells with siRNA duplexes (siE2F2 or the negative control) and miR-155 inhibitors (miR-155 or the negative control), the levels of E2F2 protein and EMT-related proteins were measured by Western blot. ( D ) E2F2 overexpression counteracted the positive proliferative effects of miR-155 in 786-O cells. ( E ) E2F2 inhibition hampered the negative proliferative function of miR-155 inhibitor in ACHN cells. ( F ) Cell migration and invasion in 786-O cells were analyzed by transwell assays. Cells were co-transfected with lentiviral particles (E2F2 or the empty vector) and miR-155 (mimic or the NC mimic). ( G ) Knockdown of E2F2 may reverse the effect of the miR-155 inhibitor on migration and invasion in ACHN cells. Data represent the mean ± SD. (* P < 0.05; ** P < 0.01; *** P < 0.001).

Article Snippet: Membranes were blocked and then incubated with primary antibodies against E2F2 (Santa Cruz, sc-632, USA), N-Cadherin (Abcam, Ab76057), E-Cadherin (Cell Signaling Technology, #3195, USA), Vimentin (Cell Signaling Technology, #5741, USA), ZEB1 (Cell Signaling Technology#3396, USA), or β-actin (ZSGB-BIO, TA-09, China) followed by incubation with horseradish peroxidase-conjugated secondary antibodies.

Techniques: Control, Plasmid Preparation, Transfection, Quantitative RT-PCR, Cotransfection, Negative Control, Western Blot, Over Expression, Inhibition, Migration, Knockdown

Journal: Oncotarget

Article Title: miR-155 regulates the proliferation and invasion of clear cell renal cell carcinoma cells by targeting E2F2

doi: 10.18632/oncotarget.7951

Figure Lengend Snippet: ( A ) Mouse tumors were obtained and dissected 4 weeks after subcutaneous injection of the transfected 786-O/miR-155 plasmid or 786-O/EV transfected cells. ( B , C ) Comparison of tumor weights and volumes between the 786-O/miR-155 and 786-O/EV groups. ( D ) Protein levels of E2F2 were immunohistochemically evaluated in the experimental and control groups. ( E ) EMT-related markers, including E-cadherin, ZEB1, and vimentin, were detected by Western blot between the groups. Data represent the mean ± SD. (* P < 0.05; ** P < 0.01; *** P < 0.001).

Article Snippet: Membranes were blocked and then incubated with primary antibodies against E2F2 (Santa Cruz, sc-632, USA), N-Cadherin (Abcam, Ab76057), E-Cadherin (Cell Signaling Technology, #3195, USA), Vimentin (Cell Signaling Technology, #5741, USA), ZEB1 (Cell Signaling Technology#3396, USA), or β-actin (ZSGB-BIO, TA-09, China) followed by incubation with horseradish peroxidase-conjugated secondary antibodies.

Techniques: Injection, Transfection, Plasmid Preparation, Comparison, Control, Western Blot

Journal: International Journal of Clinical and Experimental Pathology

Article Title: E2F2 induction in related to cell proliferation and poor prognosis in non-small cell lung carcinoma

doi:

Figure Lengend Snippet: Table 1

Article Snippet: The membranes were blocked with 5% nonfat milk and incubated with primary antibody against E2F2 (at a 1:1000 dilution, Cell signaling technology, USA) and GAPDH (1:1000, Santa Cruz, USA) at 4°C for overnight.

Techniques: Expressing