Super-Resolution Imaging Approaches for Quantifying F-Actin in Immune Cells

Abstract

Immune cells comprise a diverse set of cells that undergo a complex array of biological processes that must be tightly regulated. A key component of cellular machinery that achieves this is the cytoskeleton. Therefore, imaging and quantitatively describing the architecture and dynamics of the cytoskeleton is an important research goal. Optical microscopy is well suited to this task. Here, we review the latest in the state-of-the-art methodology for labeling the cytoskeleton, fluorescence microscopy hardware suitable for such imaging and quantitative statistical analysis software applicable to describing cytoskeletal structures. We also highlight ongoing challenges and areas for future development.

3D live imaging and phenotyping of the subcellular cytotoxicity in cancer immunotherapy using event-triggered Bessel oblique plane microscopy

“…After 40 chosen cells were imaged, they were returned to the first cell, and the above process was repeated, with each imaging cycle lasting 10 minutes (15 s/cell × 40 cells). LIS not only improved the imaging throughput but also notably reduced the phototoxicity to cells, extending the imaging window of CAR-T cells from a few minutes 21 to several hours until the endpoint was reached. The combination of CIS and LIS thereby established cross-validation for compressive visualization of the cytotoxic activities.…”

TatA and TatB generate a hydrophobic mismatch important for the function and assembly of the Tat translocon in Escherichia coli

“…Bridged-TatA assemblies likely diffuse together and therefore were not distinguishable from individual TatA assemblies within the 50 nm pixels in these fluorescence analyses ( 23 ). In contrast, METTEM resolved the 1 nm MT3-gold tags and therefore had a higher resolution than the current limit of ∼20 nm in super-resolution fluorescence techniques ( 58 ). To our knowledge, this is the first application for METTEM in localization of membrane protein associations in any organism.…”