Abstract 4229: Physical expansion of tissue microarrays for high-resolution imaging of normal and cancer samples with conventional microscopy
Authors: Bucur Octavian , Zhao Yongxin , Boyden Edward , Beck Andrew H. 
Published: July 2016

Abstract
BACKGROUND
Archival formalin fixed paraffin embedded (FFPE) tissues are an excellent and abundant source of human cancer and normal tissues, useful for cancer research and diagnosis. Moreover, FFPE tissue microarrays enable analysis of hundreds of samples from many different patients on the same slide. Super resolution microscopes have been used for cancer tissue imaging; however, these methods are expensive and require long recording times, limiting their usefulness for the cancer research community. Recently, a new approach (Expansion Microscopy) was developed to enable physical magnification and high resolution imaging of cell lines and fresh-frozen (and fixed) mouse brain with conventional microscopes (Chen F, Tillberg PW, Boyden ES, 2015, Science 347:543-548). In the current study, we developed expansion microscopy for expanding and imaging formalin fixed paraffin embedded normal and cancer human tissues, both in tissue microarrays and whole tissue slides.
METHODS
Expansion microscopy (ExM) physically magnifies tissue samples by embedding them in a dense swellable polymer, anchoring key biomolecules to the polymer mesh, and adding water to swell the polymer. ExM physically magnifies the brain tissue with nanoscale isotropy and a post-expansion measurement error of 2-4%. Proteins can also be visualized by a modified immunofluorescence assay (Chen F et al., Science, 2015).
In the present study, we optimized the ExM chemistry, labeling, and imaging methodologies to enable ExM to be used in cancer research and diagnosis, for morphological and protein imaging and analysis of both tissue microarrays and whole tissue slides on a wide variety of human tissues. Nuclei were detected by DAPI, and protein markers, including those of stroma (vimentin) and epithelium (keratins), were detected using immunofluorescence.
RESULTS and CONCLUSIONS
We developed ExM protocols to enable expansion of human normal and cancer tissues ∼4.5x in linear dimension, with a post-expansion measurement error of 5% or less. We successfully expanded normal and cancer human FFPE tissue microarrays containing over eight different tissue origins, including breast, prostate, lung, colon, pancreas and ovary. We expanded fresh frozen tissue samples of normal and cancer tissues. Our method makes possible the use of ExM on routinely collected FFPE pathology samples, enabling super resolution optical investigation of morphology and protein expression/localization in tissue microarrays with conventional fluorescent microscopy. Future applications include both large-scale retrospective and prospective studies of carcinogenesis in clinical tissue samples. * Yongxin Zhao and Octavian Bucur contributed equally; # Edward Boyden (esb@media.mit.edu) and Andrew H. Beck are corresponding authors (abeck2@bidmc.harvard.edu);
Citation Format: Octavian Bucur, Yongxin Zhao, Edward Boyden, Andrew H. Beck. Physical expansion of tissue microarrays for high-resolution imaging of normal and cancer samples with conventional microscopy. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 4229.