期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2015
卷号：112
期号：37
页码：11624-11629
DOI：10.1073/pnas.1515121112
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：SignificanceMicroscopic imaging of DNA has to rely on the use of fluorescent staining, an exogenous labeling in biological and biomedical studies, which often leads to uncertainty with respect to the quality and homogeneity of the staining. Label-free imaging of DNA will enable noninvasive visualization of live cell nuclei in both human and animals. Spontaneous Raman microspectroscopy offers label-free chemical contrast for DNA imaging; however, its slow imaging speed hampers its wide application for in vivo and dynamic studies. Here we developed a novel and simple approach with multicolor stimulated Raman scattering microscopy to evaluate rapid DNA imaging, which can be applied to both in vivo DNA dynamic studies and instant label-free human skin cancer diagnosis. Label-free DNA imaging is highly desirable in biology and medicine to perform live imaging without affecting cell function and to obtain instant histological tissue examination during surgical procedures. Here we show a label-free DNA imaging method with stimulated Raman scattering (SRS) microscopy for visualization of the cell nuclei in live animals and intact fresh human tissues with subcellular resolution. Relying on the distinct Raman spectral features of the carbon-hydrogen bonds in DNA, the distribution of DNA is retrieved from the strong background of proteins and lipids by linear decomposition of SRS images at three optimally selected Raman shifts. Based on changes on DNA condensation in the nucleus, we were able to capture chromosome dynamics during cell division both in vitro and in vivo. We tracked mouse skin cell proliferation, induced by drug treatment, through in vivo counting of the mitotic rate. Furthermore, we demonstrated a label-free histology method for human skin cancer diagnosis that provides comparable results to other conventional tissue staining methods such as H&E. Our approach exhibits higher sensitivity than SRS imaging of DNA in the fingerprint spectral region. Compared with spontaneous Raman imaging of DNA, our approach is three orders of magnitude faster, allowing both chromatin dynamic studies and label-free optical histology in real time.
