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CIL:12954*  Cite 

Immunoelectron microscopy of mammary gland tissue from lactating rats investigating potential structural interactions between mature lipid droplets and elements of the trans-Golgi network, identified by TGN38 (a transmembrane trans-Golgi protein; larger gold particles) and GOLPH3 (a trans-Golgi associated protein that can also be associated with mitochondria; smaller gold particles) staining. Note that trans‐Golgi cisternae in lactating mammary cells contain large amounts of casein and other milk proteins and are grossly expanded due to the osmolarity of the secreted fluid.

Technical Details

Tissue was processed for immunoelectron microscopy using a modified Tokuyasu method. Briefly, minced tissue was fixed in 4% paraformaldehyde containing 5% sucrose and 100 mM HEPES and infiltrated with PBS containing 2.1 M sucrose over 10 h, with repeated solution changes. Fixed tissue was transferred to an aluminum cryosectioning stub (Ted Pella, Inc., Redding, CA) and immediately frozen in liquid nitrogen. Semithin (90 nm) cryosections were cut at -110C with an UltraCut UCT/FCS cryomicrotome (Leica), using a diamond knife (Diatome) and transferred to a Formvar-coated, carbon-coated, glow-discharged 100-mesh copper-rhodium electron microscopy grid. Following blocking of nonspecific antibody binding sites with 10% calf serum in PBS, the sections were labeled by sequential incubation with antibodies to GOLPH3 (rabbit polyclonal GMx33) and TGN38 (mouse monoclonal 2F7) and colloidal gold-conjugated secondary antibodies (10 nm anti-rabbit, 15 nm anti-mouse; Ted Pella Inc., Redding, CA) and then negatively stained and embedded with 1% uranyl acetate, 1% methylcellulose in distilled water. Samples were viewed in a Philips CM10 electron microscope, and images were collected digitally. Magnification 39,000 X. See Ladinsky and Howell (2007) for more information on methods used.

Biological Sources
NCBI Organism Classification
Rattus rattus
Cell Type
mammary alveolar cell
glandular epithelial cell
milk secreting cell
Cellular Component
smooth endoplasmic reticulum
lipid particle
trans-Golgi network
Biological Context
Biological Process
lipid storage
Molecular Function
protein binding
Mark Ladinsky
Kathryn E. Howell
Ladinsky, MS and KE Howell (2007)
Digital Object Identifier (DOI)
Archival Resource Key (ARK)
Grouping This image is part of a group.
Image Type
recorded image
Image Mode
transmission electron microscopy (TEM)
illumination by electrons
detection of electrons
Parameters Imaged
electron density
Source of Contrast
differences in adsorption or binding of stain
distribution of a specific protein
Visualization Methods
uranyl salt
primary antibody plus labeled secondary antibody
Processing History
unprocessed raw data
Data Qualifiers
raw, unprocessed data
suitable for spatial measurements
Sample Preparation
cryostat-sectioned tissue
formaldehyde fixed tissue
embedded tissue
methylcellulose embedded
Relation To Intact Cell
sectioned tissue
Spatial Axis Image Size Pixel Size
X 2716px 28.2222µm
Y 3248px 28.2222µm