Journal:

Article Title: Phospholipase D2 Is Localized to the Rims of the Golgi Apparatus in Mammalian Cells

doi: 10.1091/mbc.02-04-0059

Figure Lengend Snippet: PLD2 localization with different organelles in rat NRK cells. Cells were prepared for immunofluorescence microscopy and incubated with rabbit PLD2 antibody PLD2-27 (A, D, G, and J) and costained with monoclonal antibodies to the ER marker BiP (B); the transferrin receptor (E), a marker of early endosomes and the plasma membrane; and lgp120 (H), a late endosome/lysosomal marker. (K) Cells were stained with caveolin-1, a marker for caveoli; the arrow corresponds to caveolin-1 localized to the plasma membrane. Images were merged to determine overlap between PLD2 (red) and the respective marker proteins (green; C, F, I, and L). Areas of maximal overlap are yellow. (L) Inset, enlargement of the region of overlap between PLD2 and caveolin-1 in the perinuclear Golgi apparatus. The arrow indicates areas where PLD2 puncta (red) either overlap completely with caveolin-1 (green) or are directly adjacent. Images are from projected Z-series. Bars, 10 μm.

Article Snippet: Purified monoclonal anti-rat transferrin receptor antibodies were purchased from Cedarlane Laboratories (Hornby, Ontario, Canada).

Techniques: Immunofluorescence, Microscopy, Incubation, Marker, Staining

Journal:

Article Title: Survival of Aspergillus fumigatus in Serum Involves Removal of Iron from Transferrin: the Role of Siderophores

doi: 10.1128/IAI.72.3.1402-1408.2004

Figure Lengend Snippet: Degradation of transferrin by A. fumigatus in liquid cultures. A. fumigatus was incubated in MEM containing 10% human serum (A) or 2.5 μM holotransferrin (B). Supernatants were withdrawn from the cultures after the number of hours indicated above the lanes, and the presence of transferrin was determined by Western blotting following sodium dodecyl sulfate-PAGE. Controls (lanes C) were uninoculated samples incubated for 286 h. The band underneath transferrin in panel A is another protein that cross-reacted with the polyclonal antitransferrin antibody.

Article Snippet: Gels were silver stained or transferred to polyvinylidene difluoride membranes (Bio-Rad), blocked with 5% bovine serum albumin, probed with a rabbit immunoglobulin G fraction of anti-human transferrin (1:1,000 dilution; Rockland Inc., Gilbertsville, Pa.), and treated with goat anti-rabbit horseradish peroxidase.

Techniques: Incubation, Western Blot

Journal:

Article Title: Survival of Aspergillus fumigatus in Serum Involves Removal of Iron from Transferrin: the Role of Siderophores

doi: 10.1128/IAI.72.3.1402-1408.2004

Figure Lengend Snippet: Iron removal from transferrin by A. fumigatus. A. fumigatus was cultured in MEM containing 2.5 μM purified human holotransferrin (A) or 10% human serum (B). Culture media were withdrawn, and the iron saturation of transferrin was analyzed by urea-PAGE. Transferrin was visualized by Western blotting. The numbers above the lanes represent the hours of incubation with A. fumigatus. Fe2-Tf, holotransferrin; Apo-Tf, apotransferrin.

Article Snippet: Gels were silver stained or transferred to polyvinylidene difluoride membranes (Bio-Rad), blocked with 5% bovine serum albumin, probed with a rabbit immunoglobulin G fraction of anti-human transferrin (1:1,000 dilution; Rockland Inc., Gilbertsville, Pa.), and treated with goat anti-rabbit horseradish peroxidase.

Techniques: Cell Culture, Purification, Western Blot, Incubation

Journal:

Article Title: Survival of Aspergillus fumigatus in Serum Involves Removal of Iron from Transferrin: the Role of Siderophores

doi: 10.1128/IAI.72.3.1402-1408.2004

Figure Lengend Snippet: A. fumigatus can transport iron from transferrin across a dialysis membrane. A. fumigatus was inoculated into MEM in which a dialysis bag containing holotransferrin (25 μM) was suspended. A. fumigatus was incubated for 48 h, and then transferrin was withdrawn from the dialysis bag and analyzed by urea-PAGE (lane +). An uninoculated control flask containing MEM plus transferrin in a dialysis bag also was examined (lane −). Pure holotransferrin (Fe2-Tf) and apotransferrin (Apo-Tf) standards also were run.

Article Snippet: Gels were silver stained or transferred to polyvinylidene difluoride membranes (Bio-Rad), blocked with 5% bovine serum albumin, probed with a rabbit immunoglobulin G fraction of anti-human transferrin (1:1,000 dilution; Rockland Inc., Gilbertsville, Pa.), and treated with goat anti-rabbit horseradish peroxidase.

Techniques: Incubation

Journal:

Article Title: Survival of Aspergillus fumigatus in Serum Involves Removal of Iron from Transferrin: the Role of Siderophores

doi: 10.1128/IAI.72.3.1402-1408.2004

Figure Lengend Snippet: Iron saturation of transferrin following incubation with A. fumigatus siderophores. Purified desferrisiderophores were incubated with holotransferrin (25 μM) at 37°C for 16 h. Desferriferrichrome, desferriferricrocin, and desferritriacetylfusarinine C were serially diluted to final concentrations of 5 μM (lanes 1), 50 μM (lanes 2), 500 μM (lanes 3), and 5 mM (lanes 4). Controls containing holotransferrin alone also were run (lanes 0). holo-Tf, holotransferrin; Fe-Tf, monoferric transferrin; apo-Tf, apotransferrin.

Article Snippet: Gels were silver stained or transferred to polyvinylidene difluoride membranes (Bio-Rad), blocked with 5% bovine serum albumin, probed with a rabbit immunoglobulin G fraction of anti-human transferrin (1:1,000 dilution; Rockland Inc., Gilbertsville, Pa.), and treated with goat anti-rabbit horseradish peroxidase.

Techniques: Incubation, Purification

Journal: PLoS ONE

Article Title: Comparative pharmacokinetics and pharmacodynamics of the advanced Retinol-Binding Protein 4 antagonist in dog and cynomolgus monkey

doi: 10.1371/journal.pone.0228291

Figure Lengend Snippet: . A , Western blotting confirms equal cross-reactivity of polyclonal rabbit anti-human RBP4 antibodies for dog and human RBP4. Samples (65 ng) of heterologously-expressed affinity-purified preparations of histidine-tagged dog (lane 1) and human (lane 2) RBP4 were probed with the antibodies. B , The representative immunoblot comparing RBP4 levels in 10 μl plasma samples from three dogs (lanes 2–4) and three cynomolgus monkeys (lanes 5–7). Human plasma sample (lane 1) was loaded as a reference control. Transferrin antibody was used to re-probe the Western blot to confirm equal loading of plasma samples (lower panel). C , The histogram representing pixel volumes of RBP4 bands. The bars represent pixel volume means ± S.D. of the scanned bands on the immunoblot in arbitrary units.

Article Snippet: Transferrin loading control was detected using rabbit anti-human transferrin polyclonal antibodies (1:1000, catalog number ADI-VAP-EN004-D, Enzo).

Techniques: Western Blot, Affinity Purification, Clinical Proteomics, Control

Journal: Toxics

Article Title: In Vitro Evaluation of the Effects of Cadmium on Endocytic Uptakes of Proteins into Cultured Proximal Tubule Epithelial Cells

doi: 10.3390/toxics8020024

Figure Lengend Snippet: Fluorescence imaging of endocytosed proteins in mouse proximal tubule epithelial cell (PTEC)-derived S1 cells. ( A ) S1 cells were incubated with 50 µg/mL fluorescein isothiocyanate (FITC)-albumin, Alexa-transferrin, FITC-β 2 -MG, or FITC-MT (green) for 30 min and then fixed with paraformaldehyde for immunofluorescence labeling with anti-EEA1 (red). Yellow staining demonstrates the colocalization of fluorescent-labeled proteins and early endosomes. ( B ) S1 cells were incubated with Alexa-transferrin for 1, 5, 10, 15, and 30 min. The localization of Alexa-transferrin (green) and early endosomes stained with anti-EEA1 (red) was visualized by confocal microscopy. Bars, 5 µm.

Article Snippet: Rabbit anti-transferrin receptor polyclonal antibody was purchased from Abnova (Taipei, Taiwan).

Techniques: Fluorescence, Imaging, Derivative Assay, Incubation, Immunofluorescence, Labeling, Staining, Confocal Microscopy

Journal: Toxics

Article Title: In Vitro Evaluation of the Effects of Cadmium on Endocytic Uptakes of Proteins into Cultured Proximal Tubule Epithelial Cells

doi: 10.3390/toxics8020024

Figure Lengend Snippet: Effects of Cd on the endocytosis efficiencies of albumin and transferrin into S1 cells. S1 cells were exposed to CdCl 2 for 1, 3, and 6 days and then incubated with FITC-albumin ( A ) or Alexa-transferrin ( B ) for 30 min. The endocytosis efficiencies were determined by flow cytometry and expressed as percentages of the control cells (no exposure to Cd). Data are presented as means ± SD ( n = 3–4).

Article Snippet: Rabbit anti-transferrin receptor polyclonal antibody was purchased from Abnova (Taipei, Taiwan).

Techniques: Incubation, Flow Cytometry, Control

Journal: Journal of Visualized Experiments : JoVE

Article Title: Chemically-blocked Antibody Microarray for Multiplexed High-throughput Profiling of Specific Protein Glycosylation in Complex Samples

doi: 10.3791/3791

Figure Lengend Snippet:

Article Snippet: A10 , Rabbit anti-human transferrin polyclonal antibody , Transferrin , GeneTex , GTX77130.

Techniques: Plasmid Preparation, Positive Control