Quantitative analysis of immunogold labelling for ferritin in liver from control and iron-overloaded rats.
The distribution of ferritin antigenicity in control and iron-loaded rat hepatocytes was investigated with an immunogold-ferritin antibody technique. Antibody to horse spleen ferritin showed immunoreactivity as determined by dot blotting with immunogold/silver staining with purified rat liver ferritin but not with rat haemosiderin. The initial site of ferritin degradation was studied by analysing the density of gold labelling in the cytosol and lysosomes in combination with pre-embedding acid phosphatase cytochemistry. Immunoreactive ferritin was present in the cytosol, cytosolic clusters and lysosomes of normal hepatocytes. After iron-loading, the labelling density increased over tenfold in parenchymal cell cytosol with a smaller increase in Kupffer cells. Ferritin clusters contained substantially more immunoreactive ferritin than equivalent areas of lysosomes or cytosol. Analysis of the labelling density in hepatocyte lysosomes showed that, despite a striking increase in iron content, one-quarter of the lysosomes showed less immunolabelled ferritin than the cytosol. The existence of a wide range of ferritin labelling densities in the lysosomes with a large proportion unlabelled suggests that the ferritin protein shell is not degraded at a significant rate either in the cytosol or in clusters but only after incorporation into lysosomes.
Cooper
P J
PJ
Division of Clinical Cell Biology, MRC Clinical Research Centre, Harrow, Middlesex, UK.
Iancu
T C
TC
Ward
R J
RJ
Guttridge
K M
KM
Peters
T J
TJ
D000135:Acid Phosphatase / Q000032:analysis; D000818:Animals; D003600:Cytosol / Q000032:analysis / Q000201:enzymology / Q000648:ultrastructure; D005293:Ferritins / Q000032:analysis* / Q000276:immunology / Q000378:metabolism; D007150:Immunohistochemistry / Q000379:methods*; D008099:Liver / Q000032:analysis* / Q000166:cytology / Q000201:enzymology; D008247:Lysosomes / Q000032:analysis / Q000201:enzymology / Q000648:ultrastructure; D008297:Male; D008854:Microscopy, Electron / Q000379:methods; D051381:Rats; D011919:Rats, Inbred Strains
D005293:Ferritins; D000135:Acid Phosphatase