A&P Seminar - Anna-Karin Gustavsson

The Analytical and Physical Seminar Series presents: Anna-Karin Gustavsson 
Rice
Host: Carlos Baiz
Title: Mapping cellular function using 3D single-molecule tracking and super-resolution microscopy
Refreshments served at 3:15pm
Location: WEL 2.122
 

Cellular function is governed by the molecular organization and interactions at the nanoscale. In this talk I will demonstrate our recent developments for improved 3D single-molecule tracking of dynamics and super-resolution imaging of nanoscale structures throughout mammalian cells and showcase applications of our approaches for cellular imaging. First, I will describe soTILT3D, an imaging platform that consists of a steerable, dithered, single-objective tilted light sheet for optical sectioning to reduce fluorescence background, photobleaching, and the risk of photodamaging sensitive samples, together with a novel 3D nanoprinted microfluidic chip for environmental control and for reflection of the light sheet into the sample. By combining these approaches with point spread function (PSF) engineering for nanoscale localization of individual molecules in 3D; deep learning for analysis of overlapping emitters; active 3D stabilization for drift correction and long-term imaging; and Exchange-PAINT for sequential multi-target imaging without chromatic offsets, we showcase whole-cell multi-target 3D single-molecule super-resolution imaging with improved accuracy, precision, and imaging speed. Next, I will demonstrate a versatile multimodal illumination platform that integrates the optical sectioning capabilities of light sheet illumination with uniform, flat-field epi- and TIRF illumination, resulting in more precise and accurate quantitation of single-molecule data. Finally, I will discuss how novel long axial-range double-helix PSFs offer stitching-free, 3D super-resolution imaging of whole mammalian cells, simplifying the experimental and analysis procedures for obtaining volumetric nanoscale structural information. Furthermore, we show that deep learning-based analysis drastically improves the achievable imaging speed and resolution with these PSFs. These imaging approaches are versatile and can be utilized to study molecular dynamics, nanoscale structures, and molecular mechanisms to address a broad range of chemical, biological, and biomedical questions related to cellular function and pathogenesis.