Challenge areas identified from January 2019 IBIN meeting:

 

– Computational

– Probe Development

– Super-Resolution

– 3D Bioimaging Instrumentation

– 3D Biomechanics

 

Each of these challenges is described in more detail below. Please use these as a guide for developing projects for IBIN funding.

 

Computational

 

– Develop mathematical models to interpret 3D imaging data and define collective behaviours

– Machine learning tools: Background noise removal and compensating for/tracking movement

– Quantitative analysis of shape changes in 3D volumes

– Unsupervised data acquisition – identification of features whilst imaging at different scales (AI); low resolution non-toxic to high resolution

– Developed software needs to be open source – use of online repositories (potentially through connection from IBIN hub website)

 

Probe Development

 

– Probes with long emission lifetimes to discriminate from background fluorescence. More excitation/emission wavelengths, improved QY/brightness, tuneable singlet/triplet coupling for improved blinking performance, low molecular weight and can be used in live cells

– Optogenetic probes (red/far-red) to trigger cell death/signalling in cell subpopulations

– New probes for STORM/PALM – especially for multi-colour imaging

– Need better lipid-specific dyes for live imaging and probes to image cell membrane curvature

– Need long-term probes for tracking/imaging over multiple days for 3D samples

– Need tension sensor probes that are not protein specific

– A centralised way to share probes and knowledge (website links to pre-prints and white papers)

 

Super-Resolution

 

– Need standardisation/QC and ground truth tools, calibration slides and standard samples

– Finding ways to make STORM/PALM data easier to acquire/analyse from 3D samples

– New approaches to correlate super-resolution, AFM and EM datasets from same sample

– 3D Super-resolution: deep tissue (adaptive optics, multiphoton, optogenetics)

– Imaging exosomes in situ: ways to identify populations and characterise surface markers in 3D

3D Bioimaging Instrumentation

– Instrumentation and methods for rapid accurate spectral unmixing from multiple z-slices within 3D samples (≥300um)

– Combine label-free and label multimodal, compatible with live cell/tissues – eg: Spectral imaging, fluorescence lifetime, bioluminescence (luciferase), Raman and Brillouin etc.

– Single-cell optogenetic targeting in 3D (precision); needs to be low photon dose

– Improving resolution in 3D systems; eg: adaptive optics, 2-photon light sheet

3D Biomechanics

– Analysing relationships between cells and forces combined with functional imaging of signalling

– Reproducibility of 3D models: Hydrogels to incorporate native proteins, Control pore size/crosslink/stiffness and physical properties.

– Imaging of short-term mechanical events and longer-term cell fates

– Ways to image tension between cells and nuclear morphology changes in dense 3D cultures/tissues

– Cytoskeleton tension sensors to uncouple fluidity and tension