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Optimized Negative Staining: a High-throughput Protocol for Examining Small and Asymmetric Protein Structure by Electron Microscopy
Published on: August 15, 2014
Apolipoprotein A-I and B distribution in the human cornea
F Ashraf1, D G Cogan, H S Kruth
1Clinical Branch, National Eye Institute, National Institutes of Health, Bethesda, MD 20892.
Purpose:
To determine the presence and localization of apolipoprotein A-I, a marker for high density lipoprotein, and apolipoprotein B, a marker for low density lipoprotein, in human cornea; to examine the relationship of these lipoprotein markers with areas of lipid accumulation in the cornea.
Methods:
A-I and B apolipoproteins were localized in frozen sections of human corneas with specific monoclonal antibodies using avidin-biotin immunoperoxidase labelling. Corneal lipid was colocalized with apolipoproteins by oil red 0 staining of immunostained sections.
Results:
Staining data showed that apolipoprotein B and lipid accumulated in the extracellular spaces of peripheral corneal stroma. However, their distributions were not coincident. The posterior region of peripheral corneal stroma (including Descemet's membrane) often contained lipid without immunodetectable apolipoprotein B. Unexpectedly, apolipoprotein A-I was associated with many keratocytes throughout the cornea in addition to an extracellular distribution heaviest in peripheral cornea.
Conclusions:
Lipid deposits lacking apolipoprotein B in peripheral cornea suggest that if accumulated corneal arcus lipid is derived from extracellular deposition of plasma low density lipoprotein, the low density lipoprotein is altered such that it looses its immunoreactive apolipoprotein B. The unexpected association of apolipoprotein A-I with keratocytes suggests that these cells are either taking up or synthesizing a protein sharing an immunoreactive epitope with apolipoprotein A-I.
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