Vistech iSIM

The iSIM (instant structured illumination microscopy) technology is integrated into a state-of-the-art Evident (formerly Olympus) microscope platform. It combines ultra-fast 3D imaging with low phototoxicity and provides close to a two-fold resolution enhancement compared to standard confocal or wide-field fluorescence microscopy. 

Funded by the ERC (awarded to Prof Neil Brockdorff), John Fell & EPA Cephalosporin Fund (awarded to Micron), the VT-iSIM system is fully equipped with environment control, high-NA Olympus objectives and twin Hamamatsu Orca Quest qCMOS cameras for simultaneous dual-colour acquisition with highest sensitivity. 

The system’s unique capability for fast and gentle live-cell super-resolution imaging using standard fluorophores and without time-consuming post-processing opens new possibilities for Oxford researchers to observe and understand dynamic processes within living cells and tissues in unparalleled spatio-temporal detail.

Applications

• Fast and gentle live super-resolution imaging in 2D and 3D.
• True two colour simultaneous live super-resolution imaging.
• Multi-channel super-resolution on fixed samples.

Specifications

• 405 nm, 488 nm, 561 nm and 640 nm lasers
• 100x NA 1.5 UPLAPO100XOHR Olympus objective
• Dual Hamamatsu Quest qCMOS cameras
• Temperature Control and gas control

VT-iSIM was used to show that localised clustering of autism spectrum disorder (ASD)-associated proteins NLGN3/4X at the leading edge of growth cones promoted neuritogenesis in immature human neurons by inducing growth cone enlargement and influencing actin filament organization. Ultimately suggesting these proteins may have a role in earlier ASD pathogenesis prior to their established role at neuronal synapses. The VT-iSIM enabled visualisation of these key proteins of interest at the nanoscopic level in a human stem cell derived neuronal system for the first time and provided unprecedented detail at speed to generate a substantial dataset of super-resolution images suitable for quantification and publication.

Images and case study provided by Dr Nicholas Gatford (Nuffield Department of Clinical Neurosciences)

Curd, A., Cleasby, A., Makowska, K., York, A., Shroff, H., & Peckham, M. (2015). Construction of an instant structured illumination microscope. Methods, 88, 37–47. 

York, A. G., Chandris, P., Nogare, D. D., Head, J., Wawrzusin, P., Fischer, R., Shroff, H. (2013). Instant superresolution imaging in live cells and embryos via analog image processing. Nature Methods, 10(11), 1122–1126.

Gatford, N.J., Deans, P.M., Duarte, R.R., Chennell, G., Sellers, K.J., Raval, P. and Srivastava, D.P., 2022. Neuroligin-3 and neuroligin-4X form nanoscopic clusters and regulate growth cone organization and size. Human Molecular Genetics, 31(5), 674-691