Journal: BMC Cancer

Article Title: Differences in regulation of tight junctions and cell morphology between VHL mutations from disease subtypes

doi: 10.1186/1471-2407-9-229

Figure Lengend Snippet: Knockdown of p27 disrupts VHL-mediated tight junction formation . (A) RCC10 cells stably expressing WT pVHL 30 were infected with retroviruses containing shRNA constructs directed at luciferase as control (sh Luc) or p27 (sh p27). Approximately 2 weeks post-infection, cell lysates were equally loaded and subjected to p27 and α-tubulin western blotting. (B) RCC10 WT VHL cells infected with shRNA to luciferase or p27 were grown to confluence on collagen I and photographed by digital phase microscopy (original magnification of 100×). (C) RCC10 WT VHL cells infected with shRNA to luciferase or p27 were grown to confluence on coverslips and immunostaining for ZO-1 (left panels) was performed (original magnification of 1000×). DAPI labeled nuclei of corresponding cells are also shown (right panels). (D and E) 786-O cells stably expressing pVHL 19 were infected and analyzed as in (A) and (C).

Article Snippet: Rabbit anti-ZO-1 (Mid) was from Zymed.

Techniques: Stable Transfection, Expressing, Infection, shRNA, Construct, Luciferase, Western Blot, Microscopy, Immunostaining, Labeling

Journal: BMC Cancer

Article Title: Differences in regulation of tight junctions and cell morphology between VHL mutations from disease subtypes

doi: 10.1186/1471-2407-9-229

Figure Lengend Snippet: Contrasting regulation of tight junctions among VHL mutants of different disease types in RCC10 . RCC10 cells lines (as indicated; all created by retroviral infection) were grown to confluence on coverslips and immunostaining for ZO-1 (left panels) was performed (original magnification of 1000×). DAPI labeled nuclei of corresponding cells are also shown (right panels).

Article Snippet: Rabbit anti-ZO-1 (Mid) was from Zymed.

Techniques: Infection, Immunostaining, Labeling

Journal: BMC Cancer

Article Title: Differences in regulation of tight junctions and cell morphology between VHL mutations from disease subtypes

doi: 10.1186/1471-2407-9-229

Figure Lengend Snippet: Contrasting levels of ZO-1 fragments among VHL mutants of different disease types in RCC10 . RCC10 cells lines (as indicated; all created by retroviral infection) were grown to confluence. Cell lysates were prepared and equally loaded and separated by SDS-PAGE. Western blots were performed for ZO-1 and α-tubulin. Arrows indicate ZO-1 immunoreactive proteins.

Article Snippet: Rabbit anti-ZO-1 (Mid) was from Zymed.

Techniques: Infection, SDS Page, Western Blot

Journal: BMC Cancer

Article Title: Differences in regulation of tight junctions and cell morphology between VHL mutations from disease subtypes

doi: 10.1186/1471-2407-9-229

Figure Lengend Snippet: Tight junctions are affected by levels of HIF-2α in 786-O . (A) Parental 786-O cells (786-O), 786-O cells stably transfected with VHL (VHL +), and 786-O cells infected with an empty retrovirus (pSuperRetro) or infected with HIF-2α shRNAs (HIF2α shRNA), as described in , were grown for 1 week past confluence on collagen I coated culture dishes. Cell lysates were prepared, equally loaded, and western blotted for HIF-2α, cyclin D1, p27, α-tubulin and ZO-1 (both upper and lower immunoreactive species that were seen in Figure 9). (B) Indicated cell lines that were assayed in (A) were grown to confluence on coverslips and immunostained for ZO-1 (left panels; original magnification of 1000×). DAPI labeled nuclei of corresponding cells are also shown (right panels). (C) Indicated cell lines that were assayed in (A) and (B) were grown to confluence on collagen I and photographed by digital phase microscopy (original magnification of 100×).

Article Snippet: Rabbit anti-ZO-1 (Mid) was from Zymed.

Techniques: Stable Transfection, Transfection, Infection, shRNA, Western Blot, Labeling, Microscopy

Journal: Reproductive Biology and Endocrinology

Article Title: Prolonged labour associated with lower expression of syndecan 3 and connexin 43 in human uterine tissue

doi: 10.1186/1477-7827-4-24

Figure Lengend Snippet: Box and whisker plots representing mRNA expression of Syndecan 3 and Connexin 43 in normal labour (gr. 3) and in prolonged labour (gr. 4) . A. Estimation of RT-PCR product size by 1.5% agarose gel electrophoresis. Molecular weight standards contain 10 bands ranging from 100 to 1000 bp. All amplicons had expected size. mRNA expression of (B) Syndecan 3 and (C)Connexin 43. The levels of mRNA expression were normalized to housekeeping gene 28S expression. * – statistically significant differences (p < 0.05) between the group compared to normal labour.

Article Snippet: A polyclonal antibody against Connexin 43 (71–0700 and 35–5000) and rabbit anti Syndecan-3 Mid (36–2400) were supplied by Zymed Laboratories Inc. (San Francisco, USA).

Techniques: Whisker Assay, Expressing, Reverse Transcription Polymerase Chain Reaction, Agarose Gel Electrophoresis, Molecular Weight

Journal: Reproductive Biology and Endocrinology

Article Title: Prolonged labour associated with lower expression of syndecan 3 and connexin 43 in human uterine tissue

doi: 10.1186/1477-7827-4-24

Figure Lengend Snippet: The grading of syndecan 3 staining

Article Snippet: A polyclonal antibody against Connexin 43 (71–0700 and 35–5000) and rabbit anti Syndecan-3 Mid (36–2400) were supplied by Zymed Laboratories Inc. (San Francisco, USA).

Techniques:

Journal: Reproductive Biology and Endocrinology

Article Title: Prolonged labour associated with lower expression of syndecan 3 and connexin 43 in human uterine tissue

doi: 10.1186/1477-7827-4-24

Figure Lengend Snippet: Co-localization of Connexin 43 and Syndecan 3 in uterine tissue during normal labour . Sections from uterine tissue from women in normal labour (panel A-C), patients at term (panel D-F) and patients with prolonged labour (panel G-I) were obtained and prepared as described above. A monoclonal antibody against syndecan 3 was added, followed by Alexa Fluor 633 goat anti-rabbit IgG antibody. After that, a Connexin 43 antibody followed by Alexa Fluor 488 rabbit anti-mouse IgG antibody was added. The excitation for Connexin 43 (panel A, D and G) and Syndecan 3 (panel B, E and H) are shown. The merged picture demonstrates the co-localization in the tissue (panel C, F and I).

Article Snippet: A polyclonal antibody against Connexin 43 (71–0700 and 35–5000) and rabbit anti Syndecan-3 Mid (36–2400) were supplied by Zymed Laboratories Inc. (San Francisco, USA).

Techniques:

Journal: PLoS ONE

Article Title: Defects in the Outer Limiting Membrane Are Associated with Rosette Development in the Nrl −/− Retina

doi: 10.1371/journal.pone.0032484

Figure Lengend Snippet: To analyze the integrity of the OLM in the Nrl −/− retina, sections were stained with antibodies specific for Zo-1 and beta-catenin (red), components of the OLM junctions, and S-opsin (green) to label OSs. A. In the WT retina, the Zo-1 staining forms a straight continuous line. In the Nrl −/− retina, gaps in OLM staining (arrows) can be seen that correspond spatially and temporally with the appearance of nascent rosettes and aberrant photoreceptors (arrowheads). B. Beta-catenin staining can be seen at the OLM as well as in the OPL and INL. In the Nrl −/− retina, gaps in OLM staining (arrows) can be seen that correspond with the appearance of nascent rosettes and aberrant photoreceptors (arrowheads). OLM: outer limiting membrane, ONL: outer nuclear layer, INL: inner nuclear layer. Scale bar, 20 µm.

Article Snippet: Primary antibodies were used as follows: rabbit anti-S-opsin (generated in-house and characterized previously ) diluted at 1∶1000; goat anti-S-opsin (H-17, Santa Cruz Biotechnology, Santa Cruz, CA) diluted at 1∶500; rabbit anti-recoverin (generously shared by Dr. James F. McGinnis, OUHSC ) diluted at 1∶5000; mouse monoclonal anti-BrdU (G3G4, the monoclonal antibody developed by Stephen J. Kaufman was obtained from the Developmental Studies Hybridoma Bank developed under the auspices of the NICHD and maintained by the University of Iowa, Department of Biology, Iowa City, IA) diluted at 1∶10; rabbit anti-B-catenin (C2206, Sigma-Aldrich, St. Louis, MO), diluted at 1∶1000; rabbit anti-ZO-1(Mid) (40–2200, Invitrogen, Carlsbad, CA) diluted at 1∶100; mouse monoclonal anti-ezrin (3C12, Abcam, Cambridge, MA) diluted at 1∶50; rabbit anti-mCAR-LUMIj (generously shared by Dr. Cheryl Craft, USC , , , ); rabbit anti-water channel aquaporin 4 (A5971, Sigma-Aldrich, St. Louis, MO) and rabbit anti-RDS-CT (generated in-house and characterized previously ) diluted at 1∶2000.

Techniques: Staining