Identification of cellular sections with imaging mass spectrometry following freeze fracture.
Freeze-fracture techniques have been used to maintain chemical heterogeneity of frozen-hydrated mammalian cells for static TOF-SIMS imaging. The effects the fracture plane has on scanning electron microscopy and dynamic SIMS images of cells have been studied, but the implications this preparation method has on static SIMS have not been addressed to date. Interestingly, the chemical specificity and surface sensitivity of TOF-SIMS have allowed the identification of unique sections of rat pheochromocytoma cells exposed to the sample surface during freeze fracture. Using the extensive chemical information of the fractured surface, cellular sections have been determined using TOF-SIMS images of water, sodium, potassium, hydrocarbons, phosphocholine, and DiI, a fluorescent dye that remains in the outer leaflet of the cell membrane. Higher amounts of potassium have been imaged inside a cell versus the surrounding matrix in a cross-fractured cell. In other fractures exposing the cell membrane, phosphocholine and DiI have been imaged on the outer leaflet of the cell membrane, while phosphocholine alone has been imaged on the inner leaflet. In this paper, we discuss how imaging mass spectrometry isused to uniquely distinguish three possible sections of cells obtained during freeze fracture. The identification of these sections is important in choosing cells with a region of interest, like the cell membrane, exposed to the surface for a more thorough investigation with imaging static TOF-SIMS.
Department of Chemistry, Pennsylvania State University, University Park 16802, USA.
Thomas P
TP
Roddy
Donald M
DM
Cannon
Sara G
SG
Ostrowski
Nicholas
N
Winograd
Andrew G
AG
Ewing
D000310:Adrenal Gland Neoplasms / Q000473:pathology / Q000648:ultrastructure; D000818:Animals; D003584:Cytological Techniques / Q000295:instrumentation* / Q000379:methods; D003952:Diagnostic Imaging / Q000295:instrumentation* / Q000379:methods; D005614:Freeze Fracturing; D007091:Image Processing, Computer-Assisted; D008563:Membrane Lipids; D008856:Microscopy, Fluorescence; D016716:PC12 Cells; D010673:Pheochromocytoma / Q000473:pathology / Q000648:ultrastructure; D051381:Rats; D018629:Spectrometry, Mass, Secondary Ion
D008563:Membrane Lipids