## 5 Conclusions  
The observed differences in SWIR reflectance spectra between MS patients and healthy controls are driven by pathophysiological changes in brain and muscle tissues. These differences primarily involve changes in lipid content, water absorption, and tissue scattering properties, all of which reflect central neural (neurovascular dysfunction, demyelination, and inflammation), and peripheral muscular (hypotrophy, fibrosis, and fatigue) mechanisms.  
By integrating multisite NIR spectroscopy data with clinical parameters, this work seeks to establish a framework for using portable optical spectroscopy as a complementary tool in MS management, with potential applications in therapeutic monitoring by enabling phenotype stratification and personalized therapeutic interventions. Rigorous analytical and clinical validation, including standardized protocols for device calibration, data acquisition, and processing, is essential to ensure accuracy, sensitivity, specificity, and reproducibility across diverse populations. Early engagement with regulatory agencies would facilitate qualification and define the biomarker's context of use. Integration with routine MRI could enhance diagnostic confidence, enable more frequent bedside monitoring, and guide personalized treatment adjustments. Furthermore, its potential cost-effectiveness would warrant formal economic modeling to assess healthcare impact and optimize resource utilization.  
The following supporting information can be downloaded at: [[https://www.mdpi.com/article/10.3390/app15158534/s1](https://www.mdpi.com/article/10.3390/app15158534/s1)]([https://www.mdpi.com/article/10.3390/app15158534/s1](https://www.mdpi.com/article/10.3390/app15158534/s1)), Figure S1: EMSC + GLS-W + MC pre-processed spectra: (a) MS biceps vs. Normal biceps, (b) MS triceps vs. Normal triceps, (c) MS cortical vs. Normal cortical, and (d) MS brainstem vs. Normal brainstem. Figure S2: Comparison between mean and standard deviation of raw reflectance and pre-processed (EMSC + GLS-W + MC) spectra for: Normal biceps (a), MS biceps (b), Normal triceps (c), MS triceps (d), Normal cortical (e), MS cortical (f), Normal brainstem (g), and MS brainstem (h).  
Conceptualization, A.C. and G.B.; methodology, A.C., C.M., G.B., S.S., R.G., L.M. and D.G.; software, R.G. and D.G.; validation, A.C., A.M., C.M., G.B., S.S., P.M., R.G., F.F., L.M. and L.P.; formal analysis, A.C., C.M., G.B., R.G. and D.G.; investigation, A.C., C.M., G.B., R.G., D.G. and C.T.; resources, A.C. and G.B.; data curation, A.C., C.M., G.B., R.G. and D.G.; writing--original draft preparation, A.C., C.M., G.B., R.G., C.T., L.M. and L.P.; writing--review and editing, A.C., G.B., R.G., L.M., N.L.B. and L.P.; visualization, A.C., A.M., C.M., F.F., G.B., S.S., P.M., R.G., N.L.B. and D.G.; supervision, A.C., C.T. and G.B. All authors have read and agreed to the published version of the manuscript.  
This research received no external funding.  
The study was conducted according to the guidelines of the Declaration of Helsinki and approved by the local ethics committee (Comitato Etico Lazio 2, protocol number 0167183/2018).  
Informed Consent Statement: Informed consent was obtained from all subjects involved in the study.  
The datasets generated during and/or analyzed during the study are available from the corresponding author on reasonable request.  
The authors declare no conflicts of interest.