Research Papers

Second-harmonic generation and two-photon-excited autofluorescence microscopy of cardiomyocytes: quantification of cell volume and myosin filaments

[+] Author Affiliations
Samuel J. Wallace

University of Adelaide, School of Chemistry and Physics, Adelaide, South Australia, 5005, Australia

Janna L. Morrison, Kimberley J. Botting

University of South Australia, Sansom Institute, Early Origins of Adult Health Research Group, Adelaide, South Australia, 5001, Australia and University of Adelaide, Discipline of Physiology, Adelaide, South Australia, 5005, Australia

Tak W. Kee

University of Adelaide, School of Chemistry and Physics, Adelaide, South Australia, 5005, Australia

J. Biomed. Opt. 13(6), 064018 (December 15, 2008). doi:10.1117/1.3027970
History: Received December 19, 2007; Revised September 12, 2008; Accepted September 18, 2008; Published December 15, 2008
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The ability to quantify changes in cardiomyocyte and myosin volume across gestation and in response to intrauterine insults will lead to a better understanding of the link between low birth weight and an increased risk of heart disease in adult life. We present the use of second-harmonic generation (SHG) and two-photon excitation autofluorescence (TPEF) microscopy to image unstained isolated fetal cardiomyocytes. The simultaneous collection of these two images provides a wealth of information on the morphology of cardiomyocytes. The SHG signal provides high-contrast images of myosin filaments and the TPEF signal can be used to clearly visualize cell morphology. A potential issue may arise if SHG microscopy is performed exclusively due to the lack of sensitivity to distinguish between mononucleated and binucleated cardiomyocytes. However, TPEF microscopy has the ability to efficiently separate the two types of cardiomyocytes. In addition, quantitative analysis of the SHG and TPEF images enables quantification of myosin filament level and accurate determination of cell volume. In short, we demonstrate that advanced nonlinear optical microscopy can be used to answer key physiological questions in the early origins of adult health with increased accuracy and speed compared to previously used methods.

Figures in this Article
© 2008 Society of Photo-Optical Instrumentation Engineers

Citation

Samuel J. Wallace ; Janna L. Morrison ; Kimberley J. Botting and Tak W. Kee
"Second-harmonic generation and two-photon-excited autofluorescence microscopy of cardiomyocytes: quantification of cell volume and myosin filaments", J. Biomed. Opt. 13(6), 064018 (December 15, 2008). ; http://dx.doi.org/10.1117/1.3027970


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