Rapid Sequential in Situ Multiplexing with DNA Exchange Imaging in Neuronal Cells and Tissues

• DOI: 10.1021/acs.nanolett.7b02716
• Publication Date: 2017-09-21T10:55:55Z
• Authors: Yu Wang; Johannes B. Woehr...; Noah Donoghue; Mingjie Dai; Maier S. Avendaño; Ron C. J. Schackmann; Jason J. Zoeller; Shan Shan H. Wang; Paul W. Tillberg; Demian Park; Sylvain W. Lapan; Edward S. Boyden; Joan S. Brugge; Pascal S. Kaeser; George M. Church; Sarit S. Agasti; Ralf Jungmann; Peng Yin
• Keywords: Immunostaining; Molecular Imaging; STED microscopy

To decipher the molecular mechanisms of biological function, it is critical to map composition individual cells or even more importantly tissue samples in context their environment situ. Immunofluorescence (IF) provides specific labeling for profiling. However, conventional IF methods have finite multiplexing capabilities due spectral overlap fluorophores. Various sequential imaging been developed circumvent this limit but are not widely adopted common limitation requiring multirounds slow (typically over 2 h at room temperature overnight 4 °C practice) immunostaining. We present here a practical and robust method, which we call DNA Exchange Imaging (DEI), rapid situ spectrally unlimited multiplexing. This technique overcomes speed restrictions by allowing single-round immunostaining with DNA-barcoded antibodies, followed (less than 10 min) buffer exchange fluorophore-bearing imager strands. The programmability DEI allows us apply diverse microscopy platforms (with Confocal, Exchange-SIM, Exchange-STED, Exchange-PAINT demonstrated here) multiple desired resolution scales (from ∼300 nm down sub-20 nm). optimized validated use complex samples, including primary neuron cultures sections. These results collectively suggest as versatile, platform rapid, highly multiplexed imaging, potentially enabling new applications ranging from basic science, drug discovery, clinical pathology.