Journal:

Article Title: Upregulation of CD40 Expression on Endothelial Cells Infected with Human Cytomegalovirus

doi: 10.1128/JVI.76.24.12803-12812.2002

Figure Lengend Snippet: Immunoblot analysis of CD40 in whole-cell lysates. Cell monolayers were infected with HCMV TB40E at an MOI of 1 (inf.) or mock infected (noninfected [n.i.]). Cell lysates were prepared at the indicated times and analyzed by immunoblotting. (A) CD40 protein levels were detected with a polyclonal rabbit serum directed against human CD40. (B) Blots from panel A were stripped and reprobed with a rabbit serum directed against calreticulin. (C) Signal intensities from panels A and B were densitometrically quantified; data represent the ratio of CD40 to calreticulin (y axis) versus time p.i. (x axis). (D) Protein lysates from infected or noninfected cells were treated with N-glycosidase F (+) or left untreated (−) prior to immunoblot analysis with anti-CD40 antibodies. Molecular masses are given in kilodaltons.

Article Snippet: The following monoclonal antibodies or polyclonal sera were used: mouse IgG1-fluorescein isothiocyanate (FITC)-conjugated anti-human CD54 (Calbiochem, La Jolla, Calif.), anti-human major histocompatibility complex class I (MHC-I) (BD PharMingen, San Diego, Calif.), mouse IgG1-FITC anti-human CD40 (Dianova, Hamburg, Germany), mouse IgG1-FITC anti-human CD62E (Calbiochem), antibodies G28.5 and Ro1 (anti-human CD40) (gifts from H. Engelmann, Institute for Immunology, University of Munich, Munich, Germany) ( 39 ), polyclonal rabbit anti-CD40 serum (Santa Cruz Biotechnology), rabbit antiserum against human calreticulin (Dianova), biotin-conjugated rabbit anti-mouse antibodies, and biotinylated swine anti-rabbit antibodies (Dako, Hamburg, Germany).

Techniques: Western Blot, Infection

Journal: PLoS ONE

Article Title: Searching for Cellular Partners of Hantaviral Nonstructural Protein NSs: Y2H Screening of Mouse cDNA Library and Analysis of Cellular Interactome

doi: 10.1371/journal.pone.0034307

Figure Lengend Snippet: Intersection of closest shared TULV and PUUV NSs -linked nodes. Primary nodes are in red, secondary nodes in pink. Secondary nodes that do not directly connect to two different primary nodes are in white. Metanodes are represented by pink diamonds.

Article Snippet: Samples were incubated 1 h at 37°C with rabbit polyclonal antibodies against TULV NSs-GST fusion protein and mouse monoclonal anti-ACBD3 antibody (Abnova, Taipei, Taiwan).

Techniques:

Journal: PLoS ONE

Article Title: Searching for Cellular Partners of Hantaviral Nonstructural Protein NSs: Y2H Screening of Mouse cDNA Library and Analysis of Cellular Interactome

doi: 10.1371/journal.pone.0034307

Figure Lengend Snippet: Primary cells were infected with TULV strain Lodz for 9 days and fixed on coverslips. Cells were stained for the viral NSs protein, which was seen as bright spots around perinuclear area (A), and ACBD3 protein (B). FRET assay: D pre , donor intensity before bleaching, D post , donor intensity after bleaching, A pre , acceptor intensity before bleaching, A post , acceptor intensity after bleaching, FRET eff , calculated efficiency of FRET.

Article Snippet: Samples were incubated 1 h at 37°C with rabbit polyclonal antibodies against TULV NSs-GST fusion protein and mouse monoclonal anti-ACBD3 antibody (Abnova, Taipei, Taiwan).

Techniques: Infection, Staining

Journal: PeerJ

Article Title: A comprehensive analysis of the oncogenic and prognostic role of TBC1Ds in human hepatocellular carcinoma

doi: 10.7717/peerj.17362

Figure Lengend Snippet: Representative immunohistochemistry analysis of TBC1D1, TBC1D7, TBC1D8, TBC1D9b and TBC1D25 from the human protein atlas database. And we performed immunohistochemistry experiment to verify TBC1D14 protein expression in HCC.

Article Snippet: Sections were then incubated with rabbit polyclonal antibody against TBC1D14 (all from Abclonal) at 1/200 dilution overnight at 4 °C and antibody conjugated with peroxidase (HRP; 1:500 dilution) at room temperature for 2 h, then covered by 3,3-diaminobenzidine (DAB).

Techniques: Immunohistochemistry, Expressing

Journal: PeerJ

Article Title: A comprehensive analysis of the oncogenic and prognostic role of TBC1Ds in human hepatocellular carcinoma

doi: 10.7717/peerj.17362

Figure Lengend Snippet: (A) Patients with higher TBC1D1, TBC1D7 and TBC1D9b expressions showed worse OS in HCC cohorts. (B) Patients with higher TBC1D8 and TBC1D14 expressions showed worse DFS in HCC cohorts.

Article Snippet: Sections were then incubated with rabbit polyclonal antibody against TBC1D14 (all from Abclonal) at 1/200 dilution overnight at 4 °C and antibody conjugated with peroxidase (HRP; 1:500 dilution) at room temperature for 2 h, then covered by 3,3-diaminobenzidine (DAB).

Techniques:

Journal: PeerJ

Article Title: A comprehensive analysis of the oncogenic and prognostic role of TBC1Ds in human hepatocellular carcinoma

doi: 10.7717/peerj.17362

Figure Lengend Snippet: (A) Western blotting analysis of protein levels of TBC1D8, TBC1D14 and β-actin upon knockout of TBC1D8 or TBC1D14 in Hep3B cells. (B) CCK8 assay of Hep3B cells after knockout of TBC1D8 or TBC1D14 compared to controls. (C and D) Representative images of the wound healing and EdU assays that were performed with Hep3B cells after knockout of TBC1D8 or TBC1D14 compared to controls. The width of the wound and the percentage of EdU positive cells were calculated and are shown in the right bar graph. (* P < 0.05, ** P < 0.01).

Article Snippet: Sections were then incubated with rabbit polyclonal antibody against TBC1D14 (all from Abclonal) at 1/200 dilution overnight at 4 °C and antibody conjugated with peroxidase (HRP; 1:500 dilution) at room temperature for 2 h, then covered by 3,3-diaminobenzidine (DAB).

Techniques: Western Blot, Knock-Out, CCK-8 Assay

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: Nucleic Acids Research

Article Title: Understanding the role of the Q338H MUTYH variant in oxidative damage repair

doi: 10.1093/nar/gkt130

Figure Lengend Snippet: Comparison of the DNA–glycosylase activity of recombinant wild-type and Q338H MUTYH proteins. ( A ) Representative plot of product formation under multiple turnover conditions obtained by reacting the 6-FAM–DNA substrate (10 nM) with 2 nM wild-type (circle) or Q338H MUTYH (triangle) in 10 µl reaction buffer at 37°C. Aliquots were withdrawn at different times ranging from 30 s to 60 min and processed by alkali treatment. Reaction products were analysed by electrophoresis on a 20% polyacrylamide–urea gel in 1× Tris-borate-EDTA (TBE) at 500 V for 3 h. Fluorescent bands, visualized by Typhoon 9200 Gel Imager, were quantified using the public domain NIH Image J software. The concentration of the cleaved product (nM) was plotted as a function of time and data fitted by Kaleidagraph software. The active fraction of both wild-type and Q338H MUTYH were evaluated by fitting data to . ( B ) Representative plot of product formation under single-turnover condition. DNA (2 nM) was reacted with 20 nM active MUTYH proteins in 10 µl of reaction buffer at 37°C. Data were fitted to and processed as described earlier in the text.

Article Snippet: For western blot analyses, cell lysates were loaded on SDS–7.5% polyacrylamide gels, transferred to nitrocellulose membrane and incubated overnight with a rabbit polyclonal antibody against MUTYH (dilution 1:200, Novus Biologicals, Littleton, CO, USA).

Techniques: Comparison, Activity Assay, Recombinant, Electrophoresis, Software, Concentration Assay

Journal: Nucleic Acids Research

Article Title: Understanding the role of the Q338H MUTYH variant in oxidative damage repair

doi: 10.1093/nar/gkt130

Figure Lengend Snippet: APE1 stimulation assay. Representative plot of product formation as function of time obtained by reacting the DNA substrate (10 nM) with wild-type, Q338H or G396D MUTYH proteins (5 nM) and 1 U of APE1. Reaction products were analysed on a 20% polyacrylamide–urea gel in 1 × TBE at 500 V for 3 h. Fluorescent bands, visualized by Typhoon 9200 Gel Imager, were quantified using the public domain NIH Image J software. The concentration of the cleaved product (nM) was plotted as a function of time and data fitted by Kaleidagraph software. ( A ) Wild-type MUTYH; ( B ) Q338H; and ( C ) G396D (empty and full symbols indicate reactions with and without APE1, respectively).

Article Snippet: For western blot analyses, cell lysates were loaded on SDS–7.5% polyacrylamide gels, transferred to nitrocellulose membrane and incubated overnight with a rabbit polyclonal antibody against MUTYH (dilution 1:200, Novus Biologicals, Littleton, CO, USA).

Techniques: Software, Concentration Assay

Journal: Nucleic Acids Research

Article Title: Understanding the role of the Q338H MUTYH variant in oxidative damage repair

doi: 10.1093/nar/gkt130

Figure Lengend Snippet: SPR analysis of DNA–MUTYH interaction. Serial dilutions of ( A ) wild-type MUTYH (0.47 µM to 3.67 nM) and ( B ) Q338H MUTYH (0.37 µM to 2.65 nM) were injected onto an 8-oxodG:A substrate, immobilized on a SA chip, at 30 μl/min. The association phase was allowed for 1800 s followed by a 300 s time delay, before starting the washing procedure. The response was recorded as a function of time. The derived kinetics parameters are provided in .

Article Snippet: For western blot analyses, cell lysates were loaded on SDS–7.5% polyacrylamide gels, transferred to nitrocellulose membrane and incubated overnight with a rabbit polyclonal antibody against MUTYH (dilution 1:200, Novus Biologicals, Littleton, CO, USA).

Techniques: Injection, Derivative Assay

Journal: Nucleic Acids Research

Article Title: Understanding the role of the Q338H MUTYH variant in oxidative damage repair

doi: 10.1093/nar/gkt130

Figure Lengend Snippet: MBP-pull down assay. The physical interaction between MBP–MUTYH and the RAD9 and HUS1 components of the 9–1–1 complex was revealed by immunomagnetic isolation of MBP–MUTYH by using anti-MBP beads. Pull-down proteins were separated by SDS–PAGE, blotted and analysed with MUTYH-, RAD9- and HUS1-specific antibodies. ( A ) Nuclear extracts incubated in absence (lane 1) or in presence of wild-type (lane 6) and Q338H MBP–MUTYH (lane 7) and pulled down. Lanes 3 and 4, beads-immobilized wild-type and Q338H proteins; lanes 5 and 8, 30 µg of nuclear extract. In lane 2, a molecular weight marker (M) has been visualized by merging the colorimetric scan of the same membrane by Image Lab™ software. Evaluation of relative levels of RAD9 and HUS1 bound to wild-type and Q338H MBP–MUTYH. Data are the mean ± SD of three experiments. The asterisks (*) and (**) indicate significant differences by Student’s t -test ( P < 0.05 and P < 0.01, respectively).

Article Snippet: For western blot analyses, cell lysates were loaded on SDS–7.5% polyacrylamide gels, transferred to nitrocellulose membrane and incubated overnight with a rabbit polyclonal antibody against MUTYH (dilution 1:200, Novus Biologicals, Littleton, CO, USA).

Techniques: Pull Down Assay, Isolation, SDS Page, Incubation, Molecular Weight, Marker, Membrane, Software

Journal: Nucleic Acids Research

Article Title: Understanding the role of the Q338H MUTYH variant in oxidative damage repair

doi: 10.1093/nar/gkt130

Figure Lengend Snippet: Sensitivity to KBrO 3 of Mutyh − / − MEFs complemented with wild-type (squares) and Q338H (triangles) MUTYH cDNA or without the cDNA insert (circles) . Survival was determined by clonal assays after 30 min exposure to increasing concentration of KBrO 3 . Data represent the mean ± SD of three independent determinations. Data for Mutyh −/− cells are taken from the study conducted by Maltore et al. .

Article Snippet: For western blot analyses, cell lysates were loaded on SDS–7.5% polyacrylamide gels, transferred to nitrocellulose membrane and incubated overnight with a rabbit polyclonal antibody against MUTYH (dilution 1:200, Novus Biologicals, Littleton, CO, USA).

Techniques: Concentration Assay

Journal: Nucleic Acids Research

Article Title: Understanding the role of the Q338H MUTYH variant in oxidative damage repair

doi: 10.1093/nar/gkt130

Figure Lengend Snippet: Cell cycle progression in wild-type and Q338H-expressing MEFs after exposure to KBrO 3 . ( A ) Cytofluorimetric analysis of cell cycle of Mutyh −/− MEFs expressing wild-type and Q338H cDNAs. Cells were exposed to 30 and 60 mM KBrO 3 and sampled for analysis at the indicated time points. ( B ) The panels show the percentage of cells in G 1 (black bars), S (grey bars) and G 2 (open bars) phases of the cell cycle.

Article Snippet: For western blot analyses, cell lysates were loaded on SDS–7.5% polyacrylamide gels, transferred to nitrocellulose membrane and incubated overnight with a rabbit polyclonal antibody against MUTYH (dilution 1:200, Novus Biologicals, Littleton, CO, USA).

Techniques: Expressing

Journal: Nucleic Acids Research

Article Title: Understanding the role of the Q338H MUTYH variant in oxidative damage repair

doi: 10.1093/nar/gkt130

Figure Lengend Snippet: Steady-state levels and repair kinetics of DNA 8-oxodG. ( A ) Steady-state levels of DNA 8-oxodG in Mutyh −/− (black bar), Mutyh −/− MEFs complemented with wild-type (open bar) and Q388H (grey bar) MUTYH cDNA. Data are the mean ± SD of four experiments. ( B ) Levels of DNA 8-oxodG were measured at the indicated time points in Mutyh −/− (circles) and Mutyh −/− MEFs complemented with wild-type (squares) and Q388H (triangles) MUTYH cDNA, after 30 min treatment with 20 mM KBrO 3 . Data are the mean ± SD of two experiments. P -values were calculated by Student’s t -test.

Article Snippet: For western blot analyses, cell lysates were loaded on SDS–7.5% polyacrylamide gels, transferred to nitrocellulose membrane and incubated overnight with a rabbit polyclonal antibody against MUTYH (dilution 1:200, Novus Biologicals, Littleton, CO, USA).

Techniques: