Nuclear microscopy: biomedical applications

doi:10.1016/0168-583X(93)95552-G

Nuclear Instruments and Methods in Physics Research Section B: Beam Interactions with Materials and Atoms

Volume 77, Issues 1–4, 1 May 1993, Pages 249-260

https://doi.org/10.1016/0168-583X(93)95552-G Get rights and content

Abstract

Recent developments in high energy ion beam techniques and technology have enabled the scanning proton microprobe (SPM) to make advances in biomedical research. In particular the combination of proton induced X-ray emission (PIXE) to measure the elemental concentrations of inorganic elements, Rutherford backscattering spectrometry (RBS) to characterise the organic matrix, and scanning transmission ion microscopy (STIM) to provide information on the density and structure of the sample, represents a powerful set of techniques which can be applied simultaneously to the specimen under investigation. This paper reviews briefly the biomedical work using the proton microprobe that has been carried out since the 2nd Int. Conf. on Nuclear Microprobe Technology and Applications held in Melbourne, 1990. Three recent and diverse examples of medical research are also presented from work carried out using the Oxford SPM. The first is a preliminary experiment carried out using human hair as a monitor for potential toxicity, using PIXE elemental mapping across the hair cross section to differentiate between elements contained within the hair and contamination from external sources. The second example is in the use of STIM to map individual cells in freeze-dried tissue, showing the possibility of the in situ microanalysis of cells and their extracellular environment. The third is the use of PIXE, RBS and STIM to identify and analyse the elemental constituents of neuritic plaque cores in untreated freeze-dried Alzheimer's tissue. This work resolves a current controversy by revealing an absence of aluminium levels in plaque cores at the 15 ppm level.

References (65)

J.C. Overley et al.
Nucl. Instr. and Meth.
(1983)
R.M. Sealock et al.
Nucl. Instr. and Meth.
(1983)
G.J.F. Legge et al.
Nucl. Instr. and Meth.
(1982)
F. Watt et al.
Nucl. Instr. and Meth.
(1991)
U. Lindh
Nucl. Instr. and Meth.
(1991)
P. Moretto et al.
G.E. Coote
J. Pallon et al.
M. Cholewa et al.
U. Lindh et al.
Nucl. Instr. and Meth.
(1991)

W.J. Trompetter et al.

R.W. Gauldie et al.

Tissue and Cell

(1991)

R.W. Gauldie et al.

Tissue and Cell

(1990)

M. Hult et al.

Scanning Microsc.

(1992)

G. Gadd et al.

A.J.J. Bos et al.

Nucl. Instr. and Meth.

(1984)

J.A. Cookson et al.

Phys. Med. Biol.

(1975)

M. Cholewa et al.

G.S. Bench et al.

Nucl. Instr. and Meth.

(1989)

F. Watt et al.

R.W. Jacobs et al.

Can. J. Neurol. Sci.

(1989)

C.N. Martyn et al.

Lancet

(1989)

T.P. Flaten

Environ. Geochem. and Health

(1990)

D.R.Crapper McLachlan et al.

Lancet

(1991)

B. McDonald et al.

J. Neurochem.

(1991)

N. Roos et al.

Cryopreparation of Thin Biological Specimens for Electron Microscopy: Methods and Applications

A.W. Robards et al.

Low Temperature Methods in Biological Electron Microscopy

S.A.E. Johannson et al.

PIXE — a Novel Technique for Elemental Analysis

(1988)

W.-K. Chu et al.

Backscattering Spectrometry

(1978)

F. Watt et al.

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