Fluorescence Polarization Imaging of Methylene Blue Facilitates Quantitative Detection of Thyroid Cancer in Single Cells

SIMPLE SUMMARY: Accurate diagnosis of thyroid fine-needle aspiration cytology is a significant clinical challenge. A method to detect thyroid cancer at the cellular level would be invaluable to reduce diagnostic uncertainty and improve clinical decision making. We studied the ability of confocal fluorescence polarization imaging of an exogenous fluorophore, methylene blue, to provide quantitative discrimination of cancerous cells in human samples. Our results indicate that fluorescence polarization imaging provides a reliable biomarker of thyroid cancer and holds the potential to shift the paradigm of cellular level cancer diagnosis from subjective visual assessment to objective measurement. ABSTRACT: Background: Diagnostic accuracy of the standard of care fine-needle aspiration cytology (FNAC) remains a significant problem in thyroid oncology. Therefore, a robust and accurate method for reducing uncertainty of cytopathological evaluation would be invaluable. Methods: In this double-blind study, we employed fluorescence emission and quantitative fluorescence polarization (Fpol) confocal imaging for sorting thyroid cells into benign/malignant categories. Samples were collected from malignant tumors, benign nodules, and normal thyroid epithelial tissues. Results: A total of 32 samples, including 12 from cytologically indeterminate categories, were stained using aqueous methylene blue (MB) solution, imaged, and analyzed. Fluorescence emission images yielded diagnostically relevant information on cytomorphology. Significantly higher MB Fpol was measured in thyroid cancer as compared to benign and normal cells. The results obtained from 12 indeterminate samples revealed that MB Fpol accurately differentiated benign and malignant thyroid nodules. Conclusions: The developed imaging approach holds the potential to provide an accurate and objective biomarker for thyroid cancer, improve diagnostic accuracy of cytopathology, and decrease the number of lobectomy and near-total thyroidectomy procedures.