Distribution of bovine and rabbit lens α-crystallin products by MALDI imaging mass spectrometry

PURPOSE: To develop a general tissue preparation protocol for MALDI (Matrix-Assisted Laser Desorption Ionization) imaging mass spectrometry of ocular lens tissue, and to compare the spatial distributions of α-crystallin and its modified forms in bovine and rabbit lenses. METHODS: Frozen bovine and rabbit lenses were cryosectioned equatorially at −20 °C into 12 μm-thick tissue sections. Lens sections were mounted onto conductive glass slides by ethanol soft-landing to maintain tissue integrity. An ethanol/xylene washing procedure was applied to each section before matrix application to facilitate uniform matrix crystal formation across the entire tissue section. Molecular images of both α-crystallin subunits and their modified forms were obtained from mass spectral data acquired at 100 μm steps across both whole rabbit and half bovine lens sections. RESULTS: Distinct spatial patterns for the two subunits of α-crystallin and their modified forms were observed in the rabbit and bovine lens sections. While αA-crystallin was extensively degraded in the lens core of both species, rabbit lenses exhibited a greater degree of larger molecular weight truncation products. In contrast, αB-crystallin degradation was limited in both species. Interestingly, phosphorylation of αA- and αB-crystallin was most abundant in the middle cortex of both species. CONCLUSIONS: An improved method for investigating the spatial distribution of α-crystallin in the ocular lens by MALDI imaging mass spectrometry has been developed. The localization of multiple degradation products and specific regions of α-crystallin phosphorylation in bovine and rabbit lenses gives new insight into the program of lens fiber cell differentiation and normal lens function.