**Non-Perturbative Identification and Subtyping of Amyloidosis in Human Kidney Tissue with Raman Spectroscopy and Machine Learning**

**Jeong Hee Kim 1, Chi Zhang 1, Christopher John Sperati 2, Serena M. Bagnasco <sup>3</sup> and Ishan Barman 1,4,5,\***


**Abstract:** Amyloids are proteins with characteristic beta-sheet secondary structures that display fibrillary ultrastructural configurations. They can result in pathologic lesions when deposited in human organs. Various types of amyloid protein can be routinely identified in human tissue specimens by special stains, immunolabeling, and electron microscopy, and, for certain forms of amyloidosis, mass spectrometry is required. In this study, we applied Raman spectroscopy to identify immunoglobulin light chain and amyloid A amyloidosis in human renal tissue biopsies and compared the results with a normal kidney biopsy as a control case. Raman spectra of amyloid fibrils within unstained, frozen, human kidney tissue demonstrated changes in conformation of protein secondary structures. By using t-distributed stochastic neighbor embedding (t-SNE) and density-based spatial clustering of applications with noise (DBSCAN), Raman spectroscopic data were accurately classified with respect to each amyloid type and deposition site. To the best of our knowledge, this is the first time Raman spectroscopy has been used for amyloid characterization of ex vivo human kidney tissue samples. Our approach, using Raman spectroscopy with machine learning algorithms, shows the potential for the identification of amyloid in pathologic lesions.

**Keywords:** Raman spectroscopy; machine learning; renal amyloidosis; human kidney tissue; amyloid subtyping
