Purchase this article with an account.
Jun Han, Kevin L. Schey; MALDI Tissue Imaging of Ocular Lens α-Crystallin. Invest. Ophthalmol. Vis. Sci. 2006;47(7):2990-2996. https://doi.org/10.1167/iovs.05-1529.
Download citation file:
© ARVO (1962-2015); The Authors (2016-present)
purpose. To apply MALDI (matrix-assisted laser desorption ionization) tissue imaging methods to obtaining a profile of the distribution of the lens α-crystallins and their modified forms in calf and mature bovine lenses.
methods. Frozen bovine lenses were cut equatorially at −12 °C to −20°C into 10- to 40-μm sections depending on lens age. Tissue sections were mounted onto MALDI sample plates by ethanol soft-landing to maintain tissue integrity. A two-layered matrix deposition method was used to improve mass spectral reproducibility across sections. Molecular images of the two subunits of α-crystallin and their modifications over approximately one-half of a single tissue section were reconstituted from mass spectral data sets acquired in 250-μm steps. Identification of protein truncation products and confirmation of phosphorylation distribution patterns were performed by reverse-phase liquid chromatography of soluble extracts from specific tissue regions followed by tandem mass spectrometry (LC/MS/MS).
results. Distinct distribution patterns were observed for the two subunits of α-crystallin and their modified forms. αA-crystallin showed extensive truncation across whole sections, especially in the nuclei, whereas αB-crystallin was observed to be relatively stable. Both αA-crystallin and αB-crystallin displayed the highest level of phosphorylation in the middle cortex region, a finding confirmed by LC/MS/MS analysis of dissected regions.
conclusions. A new imaging technique has been successfully applied to molecularly characterize the spatial distribution of lens proteins and their modifications in lens sections. The different distributions of α-crystallin revealed in this study provide new leads in the investigation of underlying physiological significance of the modified forms of the two α-crystallin subunits.
This PDF is available to Subscribers Only