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Description

After EGF stimulation for 4 h, Helix Pomatia Lectin (HPL) (green) in HeLa cells expressing constitutively active mutant Arf1-GFP (Arf1(Q71L)-GFP) (gray) resists redistribution from Golgi (Giantin) (red) to ER. HPL binds various glycans but the Tn antigen in particular. The Tn antigen refers to terminal α-linked N-acetyl galactosamine residues (GalNAc) linked to Ser or Thr residues. HPL indicates distribution of GalNac-Ts. HeLa cells were serum starved overnight in DME (noFBS) and then treated with human recombinant EGF (100 ng/ml; Sigma-Aldrich) for 4h. Cells were fixed for 10 min (4% paraformaldehyde) and permeabilized (0.2% Triton X-100). Primary antibody staining followed the manufacturer's instructions. Cells were subsequently stained for 15塦 min with secondary Alexa 594 for anti-Giantin. Hoechst (blue) and Alexa 647-conjugated-HPL were added during secondary antibody incubations. Cells were mounted onto glass slides using FluorSave (Merck) and imaged at room temperature using an inverted FluoView confocal microscope (model IX81; Olympus) with fluorescence excitation at 488 nm, 561 nm, and 633 nm and either a 60x objective (U Plan Super Apochromatic, UPLSAPO; NA 1.35) or 100x objective (UPLSAPO; NA 1.40) using Immersol oil. Microscope coupled with a CCD camera (model FVII). Images were acquired and processed using Olympus FV10-ASW software. Image corresponds to Fig 7B, right 2 panels in J Cell Biol. 189: 843-858. 2010. Images in Fig 7 include CIL#s 13571, 13572, 13573, 13574, 13575, 13576, 13577.

Biological Sources

NCBI Organism Classification
Homo sapiens
(human)
Cell Type
epithelial cell
cervical carcinoma
Cell Line
HeLa
Cellular Component
Golgi stack
integral to membrane
nucleus

Attribution

Names
David J. Gill
Joanne Chia
Jamie Senewiratne
Frederic Bard
Published
J Cell Biol. 189: 843-858. 2010.
Link
http://jcb-dataviewer.rupress.org/jcb/...
Pubmed
20498016

Grouping

This image is part of a group.
Data Qualifiers
raw, unprocessed data
suitable for spatial measurements

Sample Preparation

Methods
formaldehyde fixed tissue
detergent permeabilized
Relation To Intact Cell
dispersed cells in vitro

Dimensions

Spatial Axis Image Size Pixel Size
X 1024px .2070μm
Y 1024px .2070μm
Z 1 .5100μm
*CIL – Cell Image Library accession number. Please use this to reference an image.