The imaging time-of-flight secondary-ion-mass-spectrometry (TOF-SIMS) method was utilized to address the problem of cholesterol localization in rat tissues. Rat kidneys were fixed, cryoprotected by sucrose, frozen, sectioned by cryoultramicrotomy, and dried at room temperature. The samples were either covered with a thin silver layer or analyzed uncovered in an imaging TOF-SIMS instrument equipped with an Au1-3(+)-source. The yield of desorbed secondary ions for some species was up to 600-fold higher after silver coating of the samples. Reference samples of cholesterol were silver-coated and analyzed by TOF-SIMS to define significant peaks, specific for cholesterol. Such peaks were found at m/z = 386 (C27H46O+), m/z = 493 (C27H46O107Ag+), m/z = 495 (C27H46O109Ag+), m/z = 879 (C54H92O2 107Ag+), and m/z = 881 (C54H92O2 109Ag+). The silver-cationized cholesterol (493 < or = m/z < or = 495) signal was localized by imaging TOF-SIMS in the kidney sections and showed a high cholesterol content in the kidney glomeruli. A more diffuse distribution of cholesterol was also found over areas representing the cytoplasm or plasma membrane of the epithelial cells in the proximal tubules of rat kidney.
Copyright 2005 Wiley-Liss, Inc.