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David M. Anderson, Kristie M. Rose, Kevin L. Schey; MALDI Imaging Of Beta And Gamma Crystallins In Bovine And Human Lenses. Invest. Ophthalmol. Vis. Sci. 2011;52(14):4749.
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© ARVO (1962-2015); The Authors (2016-present)
MALDI imaging provides an alternative multiplex imaging modality that depends on the molecular weight of the species present in tissue sections. Thus, intact and modified proteins can be imaged in a single experiment. The purpose of this study is to develop sample preparation methods that allow distributions of beta- and gamma-crystallins and their modified forms to be imaged from bovine and human lens tissue sections.
Frozen bovine and human lens sections were obtained at 20 um thickness. Lens tissue was washed with 50% acetonitrile (ACN) and sinapinic acid matrix (50:49.9:0.1 ACN:H2O:TFA) was applied using the Labcyte Portrait 630 spotter. Images were acquired using a Bruker Autoflex III in linear mode with a spatial resolution of 300 µm. Proteins were identified by top down tandem mass spectrometry (both collision induced dissociation and electron transfer dissociation) on-line with HPLC separation of lens homogenates or directly from purified lens crystallins.
A modified sample preparation protocol allowed images to be produced of beta-and gamma- crystallins from bovine and human lens sections. Gamma-S crystallin showed an outer cortical localization while other gamma crystallins were observed in the nucleus. Partially truncated beta crystallins were distributed in an outer nuclear ring.
MALDI imaging methods have been expanded to include not only alpha-crystallin, but now beta- and gamma-crystallins in lens tissues. Images can be used to detect age-related modifications and their spatial localization.
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