CLIMB raises the clinical impact of label-free imaging

Bioimaging techniques employing labels and dyes continue to break innovative new ground, as many presentations at SPIE Photonics West demonstrated in January.

Label-free imaging technologies tackle the challenge of imaging without those exogenous agents, aiming instead to collect optical data from the unaltered tissues. This approach can potentially bring advantages for both medical diagnosis and clinical practice.

“Fluorescence microscopy uses dyes or stains or labels, targeted to specific parts of the tissue or a particular molecular receptor,” said Stephen Boppart from the University of Illinois Urbana-Champaign (UIUC).

“But that means you have to start with labels and their chemistry. There is always an uncertainty as to where that label is really going relative to the tissue, and the simple presence of a contrast agent in a natural system can disrupt biological processes. Plus many of these probes may be toxic and harmful to the cell, especially when excited to fluoresce, and regulatory approval takes time. So there are some significant disadvantages.”

A label-free approach can potentially circumvent these issues, and crucially speed up the process of translating a promising imaging modality into clinical use. That’s the ultimate goal of the Center for Label-free Imaging and Multiscale Biophotonics (CLIMB), established at UIUC to develop label-free optical and computational imaging technologies as a resource for clinicians and other investigators.

Although the inherent properties of tissue often conspire to block transmitted light, there are already several established imaging modalities able to exploit endogenous contrast mechanisms in tissues and cells. These methods can analyze optical scattering, variations in refractive index or changes in polarization, and turn those parameters into imaging data. Non-linear techniques are also part of the toolbox, as Boppart explained.

“Second harmonic generation is sensitive to collagen and elastin fibers, third harmonic generation is sensitive to refractive index variations and lipid aqueous interfaces, so those provide a structural modality,” he noted. “Coherent anti-Stokes Raman scattering and stimulated Raman scattering offer more vibrational and spectroscopic information. So nature has given us a range of molecules or structures that can, under the right circumstances, produce these endogenous signals.”

Quantitative phase imaging, which analyzes the phase shift experienced by light passing through tissue, is also an important label-free technique, partly thanks to the significant strides made in computation speed and its ability to allow both imaging and data analysis in real time. AI also plays a role, sifting information from raw data and detecting patterns that had previously seemed unavailable.

Built on the work of Gabriel Popescu

CLIMB is structured in three tiers. A trio of Technology Research and Development (TRD) projects focused on quantitative phase imaging, clinical and in-vivo imaging, and computational imaging form the core of the Center’s research. Below them, eight Collaborative Projects and a further eight Service Projects have been established.

“We like to think of it in terms of push and pull,” said Boppart. “The TRDs push the technology out, and the other projects pull the technology into their own efforts, working in different areas of medicine or biology. Some are doing microscopic cell work, others are doing animal studies, while some are focused on human studies, so they are working in many different areas. We are confident that between them we have covered the landscape, and the next step is to see how well those technologies work and get adopted.”

Outreach and training are also on the Center’s agenda, with plans for summer schools in biophotonics and the development of pathways for students from underrepresented groups in STEM.

The late Gabriel Popescu, of UIUC, an expert in quantitative phase imaging and SPIE Fellow, who worked to establish the CLIMB center. Credit: SPIE.

The TRDs are led by three UIUC faculty members: Boppart, Rohit Bhargava and Mark Anastasio. But for all involved CLIMB will always be associated with UIUC’s Gabriel Popescu, an expert in quantitative phase imaging and SPIE Fellow whose belief that the Center should be created and efforts to make it happen were key to the group’s foundation and funding. Professor Popescu died in June 2022.

“Gabi had really been the lead in our efforts, and passed away just as we were waiting for confirmation of the Center’s funding and for our work to start,” said Boppart. “We see this Centre now as a legacy, a tribute to what he started and what he envisioned.”

The label-free techniques developed at CLIMB could make a major impact in cancer treatment, enhancing the always critical determination of whether the margins of a tumor have been effectively identified. That determination often requires examination of excised tissues by pathologists elsewhere, a process potentially taking several days.

CLIMB is up for collaboration

For CLIMB the ultimate goal is for real-time assistance to the surgeon, perhaps via a hand-held probe assessing the tissues at the microscopic level during the surgery itself. Another technique developed at UIUC, simultaneous label-free autofluorescence multi-harmonic (SLAM) microscopy, has already proven its worth for imaging the alternations in tumor microenvironment that take place during chemotherapy.

A neuro vascular bundle imaged by simultaneous label-free autofluorescence multi-harmonic (SLAM) microscopy — an imaging technique developed at UIUC. Credit: CLIMB / UIUC.

Treatment of neurodegenerative diseases should also benefit from an ability to monitor the impact of therapy and treatment, with Boppart’s own research including a collaboration with pharmaceutical company GSK in the GSK Center for Optical Molecular Imaging. Drug efficacy and effectiveness of targeting are parameters that could be revealed by label-free imaging of the cells being treated.

“Gone are the days where the physician can give a patient a medication and see them two weeks later to find out how they are feeling,” commented Boppart. “We need to know what has happened at the cellular level, and as soon as possible. Did that drug find its target? How was it incorporated in the cell? Was the disease altered in its pathogenesis? That’s where using these label-free optical techniques can help to reveal the molecular basis for different pharmacotherapies.”

SLAM tumor “galaxy”: Simultaneous label-free autofluorescence multi-harmonic (SLAM) microscopy — a technique developed at UIUC, is effective for imaging alternations in a tumor that occur during chemotherapy. Credit: CLIMB / UIUC

The word Multiscale in the Center’s name reflects another goal, to use cellular image data as a means of reflecting the behavior of larger tissue areas or even an entire body. Clinical translation is closely tied to a multiscale approach, commented Boppart, and the challenge will be to use microscopic imaging as a key to larger biological and medical effects.

“This is a very interdisciplinary field,” he said. “It’s not just optical science and engineering and physics, but also biology and medicine. I think this is exactly where the new discoveries are going to come from in this century, when we use this technology to produce new tools that allow us to ask questions in biology that have not been asked before. At SPIE Photonics West we are very open to discussions with potential partners from industry and elsewhere about these new questions. We want to inform the community that we exist, that we are here to help, and ready to collaborate.”

Tim Hayes is a freelance writer based in the UK. He was previously industry editor of optics.org and Optics & Laser Europe magazine. This article originally appeared in the 2023 SPIE Photonics West Show Daily.