Journal: Journal of Integrative Agriculture

Article Title: CDH1, a Novel Surface Marker of Spermatogonial Stem Cells in Sheep Testis

doi: 10.1016/s2095-3119(13)60689-9

Figure Lengend Snippet: Fig. 1 The map of the recombinant vector for expression of His-CDH1 fusion peptide. The CDH1 ectodomain was link to the His-tag in the pET-44a(+) vector. Expression of His-CDH1 fusion peptide was driven by T7 lac promoter.

Article Snippet: After blocking with 5% skim milk for 1 h at RT, the seminiferous tubules were incubated with CDH1 polyclonal antibody and anti-PLZF antibody (Santa Cruz) overnight at 4°C.

Techniques: Recombinant, Plasmid Preparation, Expressing

Journal: Journal of Integrative Agriculture

Article Title: CDH1, a Novel Surface Marker of Spermatogonial Stem Cells in Sheep Testis

doi: 10.1016/s2095-3119(13)60689-9

Figure Lengend Snippet: Fig. 2 Analysis of the optimal expressed condition of His-CDH1 fusion peptide by SDS-PAGE. M, protein molecular weight marker; 1, control; 2-9, the sample induced by IPTG at 1, 1.5, 2, 2.5, 3, 4, 5 and 6 h, respectively.

Article Snippet: After blocking with 5% skim milk for 1 h at RT, the seminiferous tubules were incubated with CDH1 polyclonal antibody and anti-PLZF antibody (Santa Cruz) overnight at 4°C.

Techniques: SDS Page, Molecular Weight, Marker, Control

Journal: Journal of Integrative Agriculture

Article Title: CDH1, a Novel Surface Marker of Spermatogonial Stem Cells in Sheep Testis

doi: 10.1016/s2095-3119(13)60689-9

Figure Lengend Snippet: Fig. 3 Western blot analysis of the condition for purifying His- CDH1 peptide using Ni-NTA chromatography. 1, prestained protein marker; 2, the recovery fluid with lysate supernatant; 3, the recovery fluid with 20 mmol imidazol; 4-5, the recovery fluid with 100 mmol imidazol; 6-7, the recovery fluid with 250 mmol imidazol; 8-9, the recovery fluid with 500 mmol imidazol; 10, the recovery fluid with 1 000 mmol imidazol.

Article Snippet: After blocking with 5% skim milk for 1 h at RT, the seminiferous tubules were incubated with CDH1 polyclonal antibody and anti-PLZF antibody (Santa Cruz) overnight at 4°C.

Techniques: Western Blot, Chromatography, Marker

Journal: Journal of Integrative Agriculture

Article Title: CDH1, a Novel Surface Marker of Spermatogonial Stem Cells in Sheep Testis

doi: 10.1016/s2095-3119(13)60689-9

Figure Lengend Snippet: Fig. 4 Ability of the sheep CDH1 polyclonal antibodies was detected by Western blotting. Total protein was from sheep testis (T), kinedy (K), pulmo (P) and liver (L). There were three bands in 135, 120 and 80 kDa.

Article Snippet: After blocking with 5% skim milk for 1 h at RT, the seminiferous tubules were incubated with CDH1 polyclonal antibody and anti-PLZF antibody (Santa Cruz) overnight at 4°C.

Techniques: Western Blot

Journal: Journal of Integrative Agriculture

Article Title: CDH1, a Novel Surface Marker of Spermatogonial Stem Cells in Sheep Testis

doi: 10.1016/s2095-3119(13)60689-9

Figure Lengend Snippet: Fig. 5 Distribution patterns of CDH1 and PLZF in cross sections of 5-mon-old sheep testis. A, immunohistochemistry examination of PLZF expressing cells within cross sections of seminiferous tubules from 5-mon-old sheep testis. B, immunohistochemistry examination of CDH1 in the same section. C, the nuclei of the cell sections were stained by Hoechst 33342. D, merged image of A, B and C. Some PLZF- and CDH1-positive cells comprising single (diamond arrows) and paired spermatogonia (arrows) were localized at the basement membrane of the seminiferous tubules. Scale bars are 40 μm.

Article Snippet: After blocking with 5% skim milk for 1 h at RT, the seminiferous tubules were incubated with CDH1 polyclonal antibody and anti-PLZF antibody (Santa Cruz) overnight at 4°C.

Techniques: Immunohistochemistry, Expressing, Staining, Membrane

Journal: Journal of Integrative Agriculture

Article Title: CDH1, a Novel Surface Marker of Spermatogonial Stem Cells in Sheep Testis

doi: 10.1016/s2095-3119(13)60689-9

Figure Lengend Snippet: Fig. 6 Whole-mount immunohistochemistry of 5-mon-old sheep seminiferous tubules. The CDH1-positive cells were small in number, there are single cells attached the basement membrane of the seminiferous tubules (arrows), and there are paired cells attached each other (diamond arrow). Scale bar is 200 μm.

Article Snippet: After blocking with 5% skim milk for 1 h at RT, the seminiferous tubules were incubated with CDH1 polyclonal antibody and anti-PLZF antibody (Santa Cruz) overnight at 4°C.

Techniques: Immunohistochemistry, Membrane

Journal: Journal of Integrative Agriculture

Article Title: Critical role of cytochrome c1 and its cleavage in porcine reproductive and respiratory syndrome virus nonstructural protein 4-induced cell apoptosis via interaction with nsp4

doi: 10.1016/s2095-3119(17)61670-8

Figure Lengend Snippet: Fig. 1 Porcine reproductive and respiratory syndrome virus (PRRSV) nonstructural protein 4 (nsp4) interacts with cytochrome c1 (cyto.c1). A, yeast two hybrid assay. The plasmids pGBKT7-nsp4 and pGADT7-cyto.c1 were co-transformed into the yeast strain Y2HGold and selected on QDO/X/ABA plates. The top and second panels are the respective positive and negative controls. B, HEK 293T cells were transfected to express Flag-cyto.c1, or HA-nsp4 or together. At 24 h post-transfection, the cells were either subjected to direct Western blot analysis or lysed and immunoprecipitated with anti-HA antibodies. The proteins bound to sepharose beads were separated by SDS-PAGE, transferred to PVDF membranes, and probed with the antibodies to Flag and HA. C, the same as B except that nsp4 truncation mutants were used. D, MARC-145 cells were transduced with letiviruses that were expressing GFP or nsp4-GFP in the presence of 8 µg mL–1 polybrene, respectively. The cells were harvested 48 h post-transduction and cell lysates were immunoprecipitated using anti-GFP beads. The proteins bound to sepharose beads were subjected to Western blot analysis using an anti-cytochrome c1 polyclonal antibody or anti-GFP mAb. E, PRRSV nsp4 interacts with cyto.c1 in mammalian cells. Vero cells grown on coverslips in six-well plates were transfected when 60–70% to express the indicated proteins, either alone or pairwise. At 18–24 h post-transfection, the cells were fixed, permeablized, and stained with antibodies to HA and Flag, and examined by confocal microscopy. For double transfections, the merged images are shown at the right.

Article Snippet: Mouse anti-green fluorescent protein (GFP) mAb (66002-1-Ig) and rabbit anti-cytochrome c1 polyclonal antibodies (10242-1-AP) were purchased from Proteintech (Chicago, IL, USA).

Techniques: Virus, Y2H Assay, Transformation Assay, Transfection, Western Blot, Immunoprecipitation, SDS Page, Transduction, Expressing, Staining, Confocal Microscopy

Journal: Urology

Article Title: Preliminary immunohistochemical characterization of a monoclonal antibody (pro:4-216) prepared from human prostate cancer nuclear matrix proteins

doi: 10.1016/s0090-4295(97)00337-3

Figure Lengend Snippet: FIGURE 2. NMP composition of normal human prostate (A) and human prostate cancer (C) taken from a radical prostatectomy specimen from a patient with clinical/y localized prostate cancer. High resolution two-dimensional gel electrophoresis of NMP preparations were isolated and analyzed as described in Methods. Panels B and D represent Western immunoblots using PRO:4-2 16 monoclonal antibody; immunoblotting technique as described in Methods. Arrowhead represents the localization of NMP(s) with immunoreactivity to the PRO:4-2 16 mouse monoclonal anti- body generated against human prostate cancer nuclear matrix preparations. kD = molecular weight in thousands; SDS-PAGE = sodium dodecyl sulfate-polyacrylamide gel electrophoresis; pl = isoelectric point.

Article Snippet: The mouse monoclonal antibodies were UROLOGY 50 (51, 1997 801 then detected with a CY3 conjugated goat anti-mouse antibody (Jackson ImmunoResearch, West Grove, Pa) and DNA was visualized with DAPI (4,6-diamidino-2-phenylindole) at 2 /.

Techniques: Two-Dimensional Gel Electrophoresis, Electrophoresis, Isolation, Western Blot, Generated, Molecular Weight, SDS Page, Polyacrylamide Gel Electrophoresis

Journal: Urology

Article Title: Preliminary immunohistochemical characterization of a monoclonal antibody (pro:4-216) prepared from human prostate cancer nuclear matrix proteins

doi: 10.1016/s0090-4295(97)00337-3

Figure Lengend Snippet: FIGURE 3. lmmunohistochemical staining of human prostate tissue with the mouse monoclonal antibody PRO:4- 2 16. Anti-mouse IgM secondary antibody (Vector’s ABC Elite Kit). 3-3’-Diaminobenzidine tetrahydrochloride (DAB) was used as a chromogen following ethyl green counterstain. (A) Histologically normal prostate tissue (nuclei dem- onstrate an IHC intensity score of 0). (B) Histologically normal prostate tissue with the emphasis on stromal nuclei (weakly positively staining stromal nuclei represent an IHC intensity score of 1). (Cl Adenocarcinoma of the prostate from a tumor with Gleason grade 3 + 3 = 6 (IHC intensity score of 3). The cancerous glands surround a noncancerous gland. (0) A prostatic gland demonstrating PIN (positively staining nuclei represent an IHC intensity score of 1 to 2). The arrow in panel B identifies an area of prostatic stroma. The small arrows in panel C identify prostatic ade- nocarcinoma glands and the large arrow in panel C identifies a histologically normal gland found among cancerous glands. The arrow in panel D represents histologic PIN.

Article Snippet: The mouse monoclonal antibodies were UROLOGY 50 (51, 1997 801 then detected with a CY3 conjugated goat anti-mouse antibody (Jackson ImmunoResearch, West Grove, Pa) and DNA was visualized with DAPI (4,6-diamidino-2-phenylindole) at 2 /.

Techniques: Staining

Journal: British Journal of Nutrition

Article Title: Mitochondrial dysfunction in the liver of type 2 diabetic Goto–Kakizaki rats: improvement by a combination of nutrients

doi: 10.1017/s0007114511000493

Figure Lengend Snippet: Fig. 5. Apoptosis-related factors p53 and p21 in liver tissue: (a) Western blotting images of p21, p53 and a-tubulin, respectively and (b) quantitative results of p53 and p21. Quantitative values were computed as the ratios of the density of the targeted protein to a-tubulin. Results are expressed as fold change of control. Values are means, with their standard errors represented by vertical bars (n 10). Mean value was significantly different from that of the Wistar group: * P,0·05, ** P,0·01. , Wistar; , GK; , GK with nutrients; , GK with pioglitazone.

Article Snippet: Liver tissue proteins were subjected to 10 % SDS-PAGE and detected with primary antibodies against p53 (1:1000, Mouse Ab no. 12 506; Santa Cruz Biotechnology, Santa Cruz, CA, USA), p21 (1:1000, Mouse Ab no. 2946; Cell Signaling Technology, Danvers, MA, USA) or a-tubulin (1:5000).

Techniques: Western Blot, Control

Journal: British Journal of Nutrition

Article Title: Maslinic acid inhibits the metastatic capacity of DU145 human prostate cancer cells: possible mediation via hypoxia-inducible factor-1α signalling

doi: 10.1017/s0007114512000967

Figure Lengend Snippet: Fig. 7. Maslinic acid reduces the activation of Akt and extracellular signal-related kinase (ERK) in DU145 cells. (A) Serum-deprived cells were incubated with 0– 25 mM-maslinic acid for 6 h, and cell lysates were prepared with or without a 15 min epidermal growth factor (EGF) stimulation. Serum-deprived cells were incu- bated with (B) 0–20 mM-LY294002 or (C) 0–40 mM-PD98059 for 6 h and lysed without stimulation or after 15 min of EGF stimulation to determine phospho (p)-Akt or p-ERK1/2 levels. To identify hypoxia-inducible factor-1a (HIF-1a), serum-deprived cells were incubated with (B) 0–20 mM-LY294002 or (C) 0–40 mM-PD98059 in the absence or presence of EGF for 6 h. Total cell lysates were subjected to Western blotting. Photographs of chemiluminescent detection of the blots, which are representative of three independent experiments, are shown. The relative abundance of each band was estimated by densitometric scanning of the exposed films, and the expression levels were normalised to those of b-actin. To determine vascular endothelial growth factor (VEGF) concentrations, serum-deprived cells were incubated with (B) 0–20 mM-LY294002 or (C) 0–40 mM-PD98059 in serum-free media with or without 10 mg/l EGF for 18 h. The 18 h conditioned media were concentrated and subjected to Western blotting with VEGF antibody. The volumes of media loaded onto the gel were adjusted for equivalent protein concen- trations. Photographs of chemiluminescent detection of the blots, which are representative of three independent experiments, are shown. The relative abundance of each band was quantified by densitometric scanning of the exposed films. The adjusted means (n 3) of each band with their standard errors are shown above each blot. a,b,c,d Mean values with unlike letters were significantly different (P , 0·05). Mr, molecular weight.

Article Snippet: The reagents used were as follows: maslinic acid (Cayman Chemical); antibodies against uPAR, TIMP-1, TIMP-2, intercellular adhesion molecule (ICAM), vascular cell adhesion molecule (VCAM) and VEGF (Santa Cruz Biotechnology); antibodies against Akt, phospho-Akt, ERK1/2 and phosphoERK1/2 (Cell Signaling Technology); anti-uPA antibody (Calbiochem); anti-HIF-1a, anti-E-cadherin and Matrigele Matrix (BD Biosciences); epidermal growth factor (EGF; R&D Systems); an adhesion assay kit (Chemicon International); transwell filters (Costar).

Techniques: Activation Assay, Incubation, Western Blot, Expressing, Molecular Weight

Journal: The Kaohsiung Journal of Medical Sciences

Article Title: Expression of Signal Transducer and Activator of Transcription 3 and Suppressor of Cytokine Signaling 3 in Urothelial Carcinoma

doi: 10.1016/s1607-551x(09)70569-8

Figure Lengend Snippet: Figure 1. Immunohistochemical staining of phosphorylated signal transducer and activator of transcription 3 (p-STAT3) Tyr705 and cytokine signaling 3 (SOCS3) in urothelial carcinomas. Nuclear staining of p-STAT3 (Tyr705): (A) low intensity grade (insert in A is normal urothelium); (B) high intensity grade. Cytoplasmic staining of SOCS3: (C) low intensity grade; (D) high intensity grade. Original magnification, 400×.

Article Snippet: The membranes were blocked with 5% non-fat dried milk in Tris-buffered Saline (pH 7.4) with Tween-20, and then incubated with either anti-phospho-STAT3 (Tyr705) (1:1,000 dilution; Cell Signaling Technology) or SOCS3 rabbit polyclonal primary antibody (Santa Cruz Biotechnology Inc.).

Techniques: Immunohistochemical staining, Staining

Journal: The Kaohsiung Journal of Medical Sciences

Article Title: Expression of Signal Transducer and Activator of Transcription 3 and Suppressor of Cytokine Signaling 3 in Urothelial Carcinoma

doi: 10.1016/s1607-551x(09)70569-8

Figure Lengend Snippet: Figure 2. Western blotting of phosphorylated signal transducer and activator of transcription 3 (p-STAT3) Tyr705 and cytokine signaling 3 (SOCS3) protein in low-grade and high-grade UCs. p-STAT3 (Tyr705) expression was low and high in low- and high-grade UCs, respectively. Lane 1 = low-grade UC; lane 2=high-grade UC. However, the expression of SOCS3 was simi- lar in low- and high-grade UCs. GADPH protein expression was used as an internal control.

Article Snippet: The membranes were blocked with 5% non-fat dried milk in Tris-buffered Saline (pH 7.4) with Tween-20, and then incubated with either anti-phospho-STAT3 (Tyr705) (1:1,000 dilution; Cell Signaling Technology) or SOCS3 rabbit polyclonal primary antibody (Santa Cruz Biotechnology Inc.).

Techniques: Western Blot, Expressing, Control