Noninvasive, label-free tissue imaging offers less alteration, more information

Developed in the 1930s, phase contrast microscopy has allowed scientists to visualize cells in beautiful detail, using nothing but tricks of the light to elegantly maximize contrast between sample and medium. This microscope, which earned its inventor Frits Zernike the Nobel Prize in Physics in 1953, set the stage for a new phase of cell microscopy: noninvasive and label-free optical imaging. 

At St. Jude, Francisco Robles, PhD, Department of Imaging Sciences, is developing next-generation tools to reveal fundamental properties of cell biology while relying solely on the inherent and emergent properties of light and their interaction with matter.

Seeing deeper without leaving a mark 

As light passes through a biological sample, it is absorbed, scattered, refracted and reflected differently by various cellular components. These interactions can be measured and converted into images that reveal tissue architecture and composition. Though fraught with complexity, noninvasive, label-free optical imaging preserves cells and tissues in their natural state without even the minor perturbation of stains, labels or dyes used in traditional microscopy methods to paint cellular compartments and biomolecules. 

While phase contrast microscopy has traditionally lacked the molecular sensitivity and three-dimensional (3D) imaging capabilities of these traditional approaches, recent technological advances by Robles have narrowed the gap, uniquely positioning his program to uncover previously unseen biological processes and support earlier disease detection.

“The complete blood count is the world’s most common medical test, but requires multiple reagents and routine calibration, making it labor-intensive. And, in the event of abnormal results, blood smears, staining and microscopic review are also required,” Robles explained. “We asked: ‘Why not skip those steps entirely?’ By directly illuminating blood samples with UV light and capturing label-free images, we can perform a complete blood count in three minutes with a compact, automated system that produces stain-free images comparable to traditional stained microscopy.”

To reach this point, however, Robles has pioneered the development of several label-free optical imaging technologies, with a particular focus on two emerging technologies each with their own hurdles to overcome: deep ultraviolet (UV) microscopy and 3D quantitative phase microscopy of thick tissues. 

Deep UV microscopy challenges established analytical techniques

“Much of our work involved challenging the assumption that UV imaging was simply not feasible for live-cell studies,” Robles said. “UV light can kill live cells, but we found that you can precisely determine the dosage at which cells begin to experience problems.” In a 2024 paper published in Biomedical Optics Express, Robles and his team found that cells could be imaged for days if the dose is properly fractionated and the experiment designed appropriately.

Robles has since demonstrated the broad capability of deep UV microscopy:

• A 2025 paper published in Laboratory Investigation showed how deep UV microscopy could assess bone marrow aspiration for diagnostic purposes as well as, or better than, standard hematopathologist inspection protocols.
• A 2026 paper published in BME Frontiers showed how the technique was functionally comparable to flow cytometry for high-throughput immune cell characterization.

“The beauty of these systems is in their simplicity,” Robles said. “You need an objective lens, a tube lens, a camera and a roughly $35 LED. The whole system we’re trying to manufacture would cost around $5,000.”

Quantitative phase microscopy brings noninvasive, real-time imaging to the clinic

While hugely promising for excised cells and tissue samples such as biopsies, deep UV microscopy is not feasible for clinical imaging (in patients). This is where quantitative phase microscopy has demonstrated huge potential. This method is built on generations of advancements in phase contrast microscopy and holography, an imaging technique that records and reconstructs the full light field reflected or transmitted by an object, i.e., a hologram. 

Researchers have found that a cell’s refractive index, which describes how much it bends light, is directly related to its dry mass. This allows them to measure how material moves within cells and study other important aspects of cell behavior. By building an extremely stable optical system and eliminating vibrations, quantitative phase microscopy can detect changes in light equivalent to less than a billionth of a meter, making it an exceptionally sensitive imaging tool.

This approach has similar limitations on thickness as deep UV microscopy; however, the Robles lab has developed a method that allows quantitative phase imaging to work in thick tissues while also continuously providing three-dimensional information. 

Using this technology, which is called quantitative oblique back-illumination microscopy, processes within the body can be imaged in real time: 

• A study published in 2026 in Biomedical Optics Express validated the technique’s potential as a tool for brain tumor imaging.
• In a study published in 2026 in Optica, T-cell behavior was evaluated in real time as they attacked glioblastoma cells within patient-derived tumor organoids.

“It was exactly the behavior we would expect biologically, but now we’re observing and measuring it for the first time. That means this can be used as a metric to evaluate the feasibility of different treatments, including CAR T–cell therapy,” Robles said. “Clinicians can take a tumor sample from a patient, grow an organoid, test different treatment combinations and determine which therapy appears most effective.”

A bright future for noninvasive imaging

The Robles lab is continuing to push the boundaries of these and other imaging technologies, going deeper in tissue, making them even more affordable, increasing image quality and integrating this system into handheld devices, such as the Deep-ultrAviolet ptychogRaphic pockeT-scope (DART), published in eLight, in collaboration with the University of Connecticut.

“We have studies underway in cervical cancer, brain tumors and skin applications. Ultimately, we want to use this anywhere we can access tissue with a probe — particularly in head and neck applications and other surgical settings,” Robles said. “We’re also working to make these systems smaller so they can be used endoscopically. There are countless potential applications for this technology.”

The fundamental principles of label-free imaging have been established for decades, but researchers, including Robles, have now reached an intersection where technological capabilities are fully meeting the challenges that have thus far prevented its full potential from being realized. As a new day breaks on imaging sciences, the path forward for noninvasive optical imaging to revolutionize the field and make a real clinical impact has been fully illuminated.