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RESEARCH PROFILES Image and reality in 3D SIMS Beauty may be skin-deep, but the z z z (a) (b) (c) chemical information in a sample 50 50 50 runs from top to bottom. That’s 0 0 0 why researchers have been trying –50 –50 –50 to expand MS imaging techniques –100 –100 –100 from two dimensions into three. –150 –150 –150 Earlier this year, researchers dem–200 –200 –200 onstrated the promise of 3D clus–250 –250 –250 ter secondary ion MS (SIMS) by –300 –300 –300 performing the technique on in0 0 0 dividual cells (Anal. Chem. 2007, 50 100 100 50 100 50 150 150 150 79, 2199–2206). The technique 200 200 200 y y y offers good depth resolution and submicrometer lateral resolution 3D images of a trehalose–peptide film on a Si substrate. (a) An uncorrected image. (b) An image corand, of course, the very high morected for the erosion rate at each pixel, calculated by dividing the total eroded depth by the total lecular specificity that MS brings. (projectile) ion fluence at each pixel. (c) Nonlinear depth-scale calibration at each pixel provides the In this issue of Analytical best result. Chemistry (pp 5529–5539), Nicholas Winograd and Juan Cheng of each point must be determined from as a cell or a tissue slice is even more Pennsylvania State University and Anthe two AFM images, and the image is complex. dreas Wucher of the University of Duiscolor-coded with the x –y dimensions in For these experiments, the team one plane and z in the other. The color burg-Essen (Germany) explore one of chose a 300-nm trehalose film—a fairly gives information about the mass at the practical limitations of 3D cluster common material—that contained a each point. SIMS—the possibility of constructing peptide and put the film on a Si subWinograd notes that several research images that don’t quite reflect the true strate. They etched a pattern in the film teams previously performed depth-pronature of the sample. The researchers (the letters PSU, in honor of Wino+ caution that a critical step in recongrad’s institution) with a Ga beam. Ga, filing experiments in which they bomtrehalose, and the peptide have very dif- barded polymer samples with beams of structing 3D images is to account for + + Au3 or SF5 and achieved good results the fact that material may be removed at ferent molecular weights and thus dra(e.g., Anal. Chem. 2004, 76, 3199– matically different removal rates. different rates throughout the sample, 3207; 2005, 77, 911–922; 3570– “Some of the mathematics was out and they present a way to make such + + 3578). But Au3 or SF5 ion beams tend there to help us correct [the depth] corrections. to damage biological samples, and only profiles, but no one had attempted to In SIMS, a sample is bombarded a limited amount of chemical informado it in a 3D imaging modality,” says with a beam of ions, and the secondary tion can be collected, he says. Instead, Winograd. The team came up with a ions that come off the surface are anaWinograd and his colleagues bombard three-step process. First, they characterlyzed in a TOF mass spectrometer. In the sample with C6+0, which induces less the 3D version, each layer of the sample ize their intact sample in detail with damage. (The 3D cluster SIMS experiis imaged and removed. The result is es- atomic force microscopy (AFM). Next, + they move the sample into the imaging ments on single cells also used a C60 sentially a depth profile for each point mass spectrometer for study. They colbeam.) on the surface. “In our case, we collectlect an MS image of the area of interest Unfortunately, 3D SIMS analysis ed 65,536 separate depth profiles,” and then turn on the C6+0 beam to etch must be done under vacuum. TechWinograd says. “We were wondering if niques such as confocal fluorescence away a layer of material. They repeat it was possible to deal with so much this cycle until they reach the depth that microscopy and confocal Raman specdata.” troscopy can be performed in air or they want. The final step is to take anAs an added complication, a rewater. But the chemical specificity of other AFM image. searcher rarely analyzes a pure material; Collecting the data takes a few hours. MS is compelling, says Winograd. “The a more likely sample is something like a killer application is if we can reconstruct Then comes the formidable and timepolymer on a silicon substrate, in which some chemical distributions in a single consuming task of transforming the case “the removal rate of the polymer is cell or tissue,” he says. “I think we’d data. The files have to be manipulated much higher than the removal rate of have a pretty powerful contribution.” a so that the AFM format can be related silicon,” Winograd says. “You have to to the MS format. The depth scale for — Elizabeth Zubritsky take that into account.” A sample such 5506
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