[Confocal laser scanning microscopy (CLSM) for validation of non-destructive histotomography of healthy bone tissue].
OBJECTIVE
Fixation (formalin), decalcification (sections) or mechanical treatment (grinding) all bear the risk of artifacts occurring during hard-tissue histology. Because studies on the etiology of pathological changes mostly focus on subclinical lesions, artifacts can simulate early changes or even be superimposed on existing changes. The objective of this study was to determine how artifacts can be reduced.


METHODS
In confocal laser scanning microscopy (CLSM) a focused laser beam scans the surface of the specimens and penetrates into the tissue. The intensity of the remitted light is recorded. The confocal effect is due to an extremely small aperture (pin-hole), excluding light from out-of-focus planes of the sample. By stepwise movement of the object table, a tomographic series of tomographic images is obtained. Sound cortical bone samples of the lower jaw (n = 20) were studied by light microscopy and by CLSM, visualizing identical areas of a ground sectioned sample after H&E staining. Additionally, embedded and fresh blocks of tissue of the same bone sample were studied histotomographically in the CLSM.


RESULTS
(1) Light microscopic micromorphology of cortical bone can be visualized adequately in the CLSM; (2) many structures that can be visualized by light microscopy only after special staining (e.g., osteozyte processes) can be visualized by the CLSM using sample blocks without pretreatment.


CONCLUSIONS
(1) Nondestructive subsurface histotomography by CLSM totally excludes mechanical artifacts; (2) physicochemical artifacts can be handled more easily because fresh samples can be studied; (3) pseudo-three-dimensional imaging allows histological interpretation of the tissue that is equivalent to macroscopic tomographic techniques (CT, MRT).
Grötz
K A
KA
Universitätsklinik für Mund-, Kiefer- und Gesichtschirurgie, Johannes Gutenberg Universität Mainz, Germany.
Piepkorn
B
B
Bittinger
F
F
Reichert
T E
TE
Duschner
H
H
de Aguiar
E G
EG
Kann
P
P
Wagner
W
W
D016477:Artifacts; D006652:Histological Techniques / Q000295:instrumentation*; D006801:Humans; D007089:Image Enhancement / Q000295:instrumentation; D008334:Mandible / Q000033:anatomy & histology*; D018613:Microscopy, Confocal / Q000295:instrumentation*
Konfokale Laser-Scanning Mikroskopie (CLSM) Validierung der zerstörungsfreien Histotomographie gesunden Knochengewebes.