Tissue section AFM: In situ ultrastructural imaging of native biomolecules.
Conventional approaches for ultrastructural high-resolution imaging of biological specimens induce profound changes in bio-molecular structures. By combining tissue cryo-sectioning with non-destructive atomic force microscopy (AFM) imaging we have developed a methodology that may be applied by the non-specialist to both preserve and visualize bio-molecular structures (in particular extracellular matrix assemblies) in situ. This tissue section AFM technique is capable of: i) resolving nm-microm scale features of intra- and extracellular structures in tissue cryo-sections; ii) imaging the same tissue region before and after experimental interventions; iii) combining ultrastructural imaging with complimentary microscopical and micromechanical methods. Here, we employ this technique to: i) visualize the macro-molecular structures of unstained and unfixed fibrillar collagens (in skin, cartilage and intervertebral disc), elastic fibres (in aorta and lung), desmosomes (in nasal epithelium) and mitochondria (in heart); ii) quantify the ultrastructural effects of sequential collagenase digestion on a single elastic fibre; iii) correlate optical (auto fluorescent) with ultrastructural (AFM) images of aortic elastic lamellae.
Graham
Helen K
HK
Unit of Cardiac Physiology, School of Biomedicine, The University of Manchester, Manchester Academic Health Science Centre, Manchester, M13 9PT, UK.
Hodson
Nigel W
NW
Hoyland
Judith A
JA
Millward-Sadler
Sarah J
SJ
Garrod
David
D
Scothern
Anthea
A
Griffiths
Christopher E M
CE
Watson
Rachel E B
RE
Cox
Thomas R
TR
Erler
Janine T
JT
Trafford
Andrew W
AW
Sherratt
Michael J
MJ
D002356:Cartilage / Q000378:metabolism / Q000648:ultrastructure*; D003952:Diagnostic Imaging / Q000379:methods; D005109:Extracellular Matrix / Q000378:metabolism / Q000648:ultrastructure*; D016591:Histocytological Preparation Techniques / Q000379:methods; D006801:Humans; D008853:Microscopy / Q000379:methods*; D018625:Microscopy, Atomic Force / Q000379:methods*