Novel Fluorescence-based Imaging Method for Slide-free Nondestructive Primary Pathology Diagnosis

by Christos Evangelou, MSc, PhD – Medical Writer and Editor

In a recent pilot validation study, researchers at the University of California Davis Health and the University of British Columbia compared a novel imaging method called fluorescence imitating brightfield imaging (FIBI) with the standard formalin-fixed, paraffin-embedded (FFPE) hematoxylin-eosin (H&E) stained tissue section histology for primary surgical pathology diagnosis.1 The study aimed to evaluate the feasibility and accuracy of the new imaging technique as a potential alternative to traditional histopathology.

The study showed that FIBI could accurately capture the surface of fresh tissue without requiring standard processing or sectioning. Across 800 reads, the overall agreement rate between FIBI and standard H&E staining was 97.9%.1

“Our findings suggest that FIBI is a valid alternative to standard formalin-fixed, paraffin-embedded H&E-stained tissue section histology for primary surgical pathology diagnosis,” said Alexander ‘Sandy’ Borowsky, MD, Professor of Pathology and Laboratory Medicine UC Davis School of Medicine and Comprehensive Cancer Center, who is the first author of the study.

He believes that this slide-free digital pathology method can help pathologists provide accurate diagnoses using slide-free images acquired through FIBI, without the need for time-consuming and labor-intensive tissue processing and staining.

The report was published in Archives of Pathology & Laboratory Medicine.


Study Rationale: Developing a Slide-free Digital Pathology Method for Primary Pathology Diagnosis

Current histological practices are often labor-intensive and time-consuming. In addition, standard histological methods can be destructive to tissue samples, leading to tissue degradation and loss of important molecular information.

“In this study, we evaluated the diagnostic quality of a newly developed technology called fluorescence imitating brightfield imaging, or simply FIBI, which offers a slide-free alternative,” Dr. Borowsky noted.

FIBI is a technically simple, fast, and cost-effective solution that is intrinsically suitable for imaging specimens, without traditional processing, and is compatible with both fresh and fixed tissue samples. “It is also nondestructive, leaving the tissue intact and available for subsequent embedding, standard histology, and downstream molecular tests if desired,” he added.



The study used a comparative design to evaluate the ability of pathologists to evaluate direct-to-digital images and standard pathology preparations.1

“We recruited four study pathologists with a range of expertise and experience but with no prior experience using our technology,” Dr. Borowsky explained.

One hundred surgical pathology samples were obtained and digitally scanned using FIBI and then processed for standard histologic examination with conventional H&E-stained sections. Cases were presented to the four study pathologists in random order and randomized FIBI or H&E. They provided the best or most likely diagnosis without additional information or ancillary studies such as immunohistochemistry. They were also asked to comment about adequacy. After a 30-day washout period, the modality was switched (if the FIBI image was seen in the first round, the H&E was provided after the washout) for the same 100 cases, and diagnosis and adequacy were again given by the study pathologists.

FIBI employs cutting-edge imaging technology that uses fluorescence microscopy optics but immediately generates images that closely resemble those seen with standard brightfield microscopes. “FIBI involves brief (1–2 min) staining of intact tissue specimens and gently compressing them against a coverslip to get a flat imaging surface. The entire tissue area is then scanned using the FIBI device,” Dr. Borowsky described.

The novelty of this approach lies in the fact that it requires very simple optics, consisting of an LED excitation source in the blue-visible range, a conventional microscope objective, and a color camera. This innovative approach eliminates the need for slide preparation and allows for rapid digital acquisition of high-resolution images and brightfield-like results.

The final dataset consisted of 100 reference diagnoses and 800 study pathologist reads. Each study read was compared to the reference diagnosis and to the reader’s own diagnosis across both imaging modalities.


FIBI Provides High Diagnostic Accuracy Comparable to that of Conventional H&E Histology

The study findings suggest that FIBI has the potential to be a reliable alternative to standard histologic examination for primary surgical pathology diagnosis. Across 800 reads, the overall agreement rate between the two modalities was 97.9%.1 This consisted of 400 digital reads at 97.0% compared with the reference and 400 standard reads at 98.8% compared with the reference.

The concordance rate between FIBI and standard H&E histology was high. When combined, the two modalities provided a 91.8% overall concordance rate, 6.1% minor discordance rate (i.e., alternative diagnoses without clinical treatment or outcome implications), and 2.1% major discordance rate (alternative diagnoses with clinical treatment or outcome implications).


Potential Applications of FIBI

When asked about the potential applications of FIBI, Dr. Borowsky said: “This method can help pathologists with on-site, nondestructive, fast, diagnostic-quality core-needle biopsy evaluation for improved rapid on-site evaluation, as well as with rapid intraoperative surgical guidance and margin assessment, replacing frozen sections.”

The fact that FIBI is simple and fast makes it an ideal point-of-care histological method for dermatology and rapid biopsy-based diagnostics, such as endoscopy and interventional radiology settings. Implementation of FIBI in central histology laboratories and research laboratories can also lead to faster turnaround times and lower costs in research settings.

“FIBI can also enable optimized precision medicine procedures by preserving available specimen quality and quantity and therefore improved DNA and RNA yields. Another benefit we envision is to provide remote diagnosis in low-resource settings, facilitated by the simple processing and easy digital capture,” Dr. Borowsky added.


Future Work

The authors suggested that FIBI could potentially lead to faster and more accurate diagnoses, which could improve treatment planning and patient outcomes. Additionally, the ability to share images instantaneously with other pathologists in different locations could facilitate collaboration and improve the quality of patient care. However, the authors also note that larger multi-site studies are needed to validate these findings and to determine the optimal use of FIBI in clinical practice.

Commenting on their plans, Dr. Borowsky said: “We would follow this study with specific use case validations at multiple sites to provide data for regulatory approvals. We plan to validate the assessment of breast lumpectomy margins with the potential to provide intraoperative guidance. Additional use cases will include rapid biopsy diagnosis and adequacy assessment, with a vision for a larger study with over 2000 cases and 12–20 pathologists to document non-inferiority to traditional FFPE slide sectioning.”

Future studies are needed to determine the feasibility of performing most or all of the assays required to guide patient care decisions at the time of biopsy by implementing immunofluorescence studies in the same imaging session.



  1. Borowsky AD, Levenson RM, Gown AM, et al. A Pilot Validation Study Comparing Fluorescence-Imitating Brightfield Imaging, A Slide-Free Imaging Method, With Standard Formalin-Fixed, Paraffin-Embedded Hematoxylin-Eosin-Stained Tissue Section Histology for Primary Surgical Pathology Diagnosis. Arch Pathol Lab Med. May 2023. doi:10.5858/arpa.2022-0432-OA

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