Abstract
5-aminolevulinic acid (5-ALA)-based fluorescence diagnosis is now clinically applied for accurate and ultrarapid diagnosis of malignant lesions such as lymph node metastasis during surgery. 5-ALA-based diagnosis evaluates fluorescence intensity of a fluorescent metabolite of 5-ALA, protoporphyrin IX (PPIX); however, the fluorescence of PPIX is often affected by autofluorescence of tissue chromophores, such as collagen and flavins. In this study, we demonstrated PPIX fluorescence estimation with autofluorescence elimination for 5-ALA-based fluorescence diagnosis of malignant lesions by simplified and optimized multispectral imaging. We computationally optimized observation wavelength regions for the estimation of PPIX fluorescence in terms of minimizing prediction error of PPIX fluorescence intensity in the presence of typical chromophores, collagen and flavins. By using the fluorescence intensities of the optimized wavelength regions, we verified quantitative detection of PPIX fluorescence by using chemical mixtures of PPIX, flavins, and collagen. Furthermore, we demonstrated detection capability by using metastatic and non-metastatic lymph nodes of colorectal cancer patients. These results suggest the potential and usefulness of the background-free estimation method of PPIX fluorescence for 5-ALA-based fluorescence diagnosis of malignant lesions, and we expect this method to be beneficial for intraoperative and rapid cancer diagnosis.
Highlights
Fluorescence detection based on 5-aminolevulinic acid (5-ALA)-induced protoporphyrin IX (PPIX) fluorescence has recently emerged as a promising method for ultrarapid intraoperative detection of malignant lesions1–6. 5-ALA-based fluorescence detection of malignant lesions is clinically applied in brain surgery[4,7], urological surgery[8,9,10], and gastrointestinal surgery[11,12]
To optimize observation wavelength for PPIX fluorescence detection, we sought the wavelength with minimum estimation error of PPIX fluorescence intensity under the condition of varied intensity ratio of PPIX fluorescence and adjacent autofluorescence
We focused on two sources of predominant autofluorescence of lymph node metastasis, derived from flavin adenine dinucleotide (FAD) and collagen, and sought to optimize the observation wavelength in the presence of random noise
Summary
Fluorescence detection based on 5-aminolevulinic acid (5-ALA)-induced protoporphyrin IX (PPIX) fluorescence has recently emerged as a promising method for ultrarapid intraoperative detection of malignant lesions1–6. 5-ALA-based fluorescence detection of malignant lesions is clinically applied in brain surgery[4,7], urological surgery[8,9,10], and gastrointestinal surgery[11,12]. 5-ALA-based fluorescence detection of malignant lesions is clinically applied in brain surgery[4,7], urological surgery[8,9,10], and gastrointestinal surgery[11,12]. This fluorescence method is based on selective accumulation of PPIX, which is a fluorescent metabolite of 5-ALA, due to altered activity of enzymes of cancer cells, the increase in porphobilinogen deaminase activity and the reduction in ferrochelatase activity[13,14,15,16]. We demonstrated detection capacity of our method by using lymph node metastases of colorectal cancer patients
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