Ultrashort light pulse generation and applications to the study of ultrafast phenomena in the matter

The main goal of the CRIMSON project is to develop an innovative bio-photonic system for cell/tissue imaging which will be used as a research tool to understand the cellular origin of diseases. Our imaging technique will be broadband coherent Raman scattering (CRS) extended to the fingerprint region, which is crucial for chemical identification. Broadband CRS combines the following advantages:

Label-free imaging, avoiding the use of exogenous or endogenous labels.
Living cell imaging capability.
High acquisition speed, enabling the observation of dynamic cellular processes by time-lapse imaging.
High biochemical sensitivity, thanks to the capability to acquire quantitative hyperspectral images allowing the identification of molecules by their unique vibrational spectrum.
The possibility to couple these advantages with endoscopy-based in vivo imaging.
Our broadband CRS microscope will extract the maximum amount of information about the biomolecular composition of the cell, coupling it with morphological information, without altering the natural state of the cell with exogenous molecules or invasive interventions. As such, it is expected to revolutionize the study of the cellular origin of diseases.

Despite its promise as a disruptive cell/tissue imaging method, CRS microscopy is currently not used by biologists as a standard research tool and remains confined to low Technology Readiness Level (TRL) laboratory applications. This is due to several limitations of current implementations of CRS:

The laser source required for CRS is complex, expensive and needs regular alignment or maintenance, making it not suitable for biomedical research laboratories.
CRS typically measures the signal at a few selected vibrational frequencies and does not provide the rich chemical information contained in spontaneous Raman, which measures the full vibrational spectrum.
Most CRS systems work in the CH stretching region (2800-3100 cm-1), which gives stronger signals but delivers less spectroscopic information with respect to the fingerprint region (700-1800 cm-1), which has exquisite sensitivity to the chemical structure.
CRIMSON aims at overcoming these limitations, by enhancing the information content of CRS microscopy and bringing it out of the research labs, making it accessible to mainstream biomedical researchers.
Specifically, CRIMSON aims at:

Developing a compact, alignment-free and low-cost laser system for CRS based on fibre laser technology, thanks to our partner Active Fiber Systems GmbH.
Extending CRS microscopy from single-frequency to broadband operation.
Extending detection of CRS signal to the fingerprint region, which provides higher information content.
Configuring the microscope system including all the requirements for time-lapse live cell imaging.
Developing a CRS flexible endoscope imaging probe compatible with in vivo semi-invasive imaging, thanks to our partner LIGHTCORE TECHNOLOGIES.
Using the extremely rich Raman spectral information in each point of the image (hyper-spectral data cube) by artificial intelligence (AI) based methods, e.g. chemometrics, machine learning and deep learning, to perform quantitative imaging with an unprecedented level of specificity and sensitivity for the study of intra- and inter-cellular processes, thanks to our partner 3rd Place.
Combining these technological advances into a complete broadband CRS microscope for commercial exploitation, coupled to a flexible CRS endoscope for future in vivo imaging, thanks to our partner CRIL.
The CRS microscope/endoscope developed in CRIMSON will offer a significant gain in the understanding of inter- and/or intra-cellular processes and it will strengthen Europe’s industrial position in the biophotonics-related market for microscopes and research and development tools.

We will validate the new imaging system using three case studies focused on cancer, selected as a paradigmatic example of the complexity and heterogeneity of human diseases with a clear cellular origin. In detail, three specific processes will be considered:

Autophagic processes, used by cells to recycle their content in order to boost their metabolism or to respond to external stress, which may be involved in cancer progression, thanks to our partner Institut National de la Santé Et de la Recherche Médicale.
The interaction between cancer and immune cells that occurs when the immune system attempts to target and attack cancer cells, which is of great relevance for immune-oncology, thanks to our partner Jena University Hospital.
Senescence, a stable cell-growth arrest involved in tumour reversion/progression. This process entails an extensive set of intra and extracellular changes that normally must be visualized by multiple approaches, thanks to our partner Istituto Nazionale dei Tumori.
The challenge of CRIMSON will be to demonstrate the capability of the CRS microscope to image and to objectively describe such complex morphological and biomolecular changes in real time.