<h3>Objective:</h3> To examine the associations between serum protein biomarker profiles and patient-reported disability in people with Multiple Sclerosis (pwMS). <h3>Background:</h3> Biomarkers could inform disease worsening and severity in pwMS. Few studies have examined blood biomarkers informative of patient-reported outcome (PRO) of disability in pwMS. Leveraging the Proximity Extension Assay methodology on the Olink™ platform, prior efforts identified a custom serum-based proteomic panel, comprising 19 proteins associated with inflammatory Multiple Sclerosis (MS) disease activity. <h3>Design/Methods:</h3> In this multicenter cross-sectional study we included 431 adults (mean age 49 years, 81% women, 94% non-Hispanic White) with MS between 2017 and 2020. For exposure, we included 19 serum protein biomarkers potentially associated with MS inflammatory disease activity and 7 key clinical factors (age at sample collection, sex, race/ethnicity, disease subtype, disease duration, disease-modifying treatment, and time interval between sample collection and closest PRO assessment). Using 4 machine learning approaches (Least Absolute Shrinkage and Selection Operator [LASSO] regression, Random Forest, XGBoost, and Support-Vector Machines, we examined model performance in predicting Patient Determined Disease Steps (PDDS) and Patient-Reported Outcomes Measurement Information System (PROMIS) physical function as outcomes. <h3>Results:</h3> Using binary outcomes, models comprising both routine clinical factors and the 19 proteins as features consistently outperformed base models (containing clinical features alone) in predicting severe (PDDS≥4, PROMIS<35) versus mild/moderate (PDDS<4, PROMIS≥35) disability for all machine learning approaches, with LASSO displaying the best area under the curve (AUC<sub>PDDS</sub>=0.91, AUC<sub>PROMIS</sub>=0.90). Using continuous outcomes, LASSO models with combined clinical and 19 proteins as features (R<sup>2</sup><sub>PDDS</sub>=0.31, R<sup>2</sup><sub>PROMIS</sub>=0.35) again outperformed base models. The four LASSO models with combined clinical and protein features shared 2 clinical features (disease subtype, disease duration) and 4 protein biomarkers (CDCP1, IL-12B, NEFL, PRTG). <h3>Conclusions:</h3> Serum protein biomarker profiles have potential clinical utility beyond clinical profile or single protein in predicting real-world MS disability status. <b>Disclosure:</b> Dr. Zhu has nothing to disclose. Miss Chen has nothing to disclose. Dr. Zhang has nothing to disclose. Miss Walker has nothing to disclose. Miss Venkatesh has received research support from Foundation of the Consortium of Multiple Sclerosis. Miss Venkatesh has received research support from NIH. Mr. Zhang has received personal compensation for serving as an employee of Octave Bioscience Inc. Mr. Qureshi has received personal compensation for serving as an employee of Octave Bioscience. Ms. Hoyt has nothing to disclose. The institution of Dr. Foley has received personal compensation in the range of $500-$4,999 for serving as a Consultant for Octave. Dr. Foley has received personal compensation in the range of $5,000-$9,999 for serving on a Scientific Advisory or Data Safety Monitoring board for Biogen. The institution of Dr. Foley has received personal compensation in the range of $500-$4,999 for serving on a Scientific Advisory or Data Safety Monitoring board for TG Therapeutics . The institution of Dr. Foley has received personal compensation in the range of $500-$4,999 for serving on a Scientific Advisory or Data Safety Monitoring board for Horizon Therapeutics. The institution of Dr. Foley has received personal compensation in the range of $500-$4,999 for serving on a Scientific Advisory or Data Safety Monitoring board for Sandoz. Dr. Foley has received personal compensation in the range of $10,000-$49,999 for serving on a Speakers Bureau for Biogen. Dr. Foley has stock in InterPRO Bioscience. The institution of Dr. Foley has received research support from Biogen. The institution of Dr. Foley has received research support from Novartis. The institution of Dr. Foley has received research support from Octave. The institution of Dr. Foley has received research support from Genentech. The institution of Dr. Foley has received research support from Imstem. The institution of Dr. Foley has received research support from Aegir. Dr. Xia has received personal compensation in the range of $5,000-$9,999 for serving as a Consultant for Genentech/Roche. Dr. Xia has received personal compensation in the range of $500-$4,999 for serving as a Consultant for Neurogene. Dr. Xia has received personal compensation in the range of $500-$4,999 for serving on a Scientific Advisory or Data Safety Monitoring board for Genentech/Roche. The institution of Dr. Xia has received research support from National Institute of Health. The institution of Dr. Xia has received research support from Octave Biosciences. The institution of Dr. Xia has received research support from Department of Defense.