Using reference intervals to improve interpretation of youth sport-related concussion biomarkers using a research platform.

Youth sports-related concussion (SRC) is a major health concern. Approximately 67% of Canadian youth participate in sports and recreation, and 93% of head injuries experienced in sports and recreation are concussions. SRCs are challenging to diagnose because symptoms are often subtle, which can affect the reliability of clinical diagnostic methods that are often subjective. As undiagnosed concussions can potentially lead to unfavourable neurological implications, objective diagnostic tools have the potential to improve diagnosis and inform prognosis. Recent research has highlighted the potential utility of plasma neurofilament light (NfL) and glial fibrillary acidic protein (GFAP) for SRC diagnosis, offering a quantitative and cost-effective diagnostic method. However, a major gap in the literature is understanding whether the magnitude of change in NfL and GFAP levels post-SRC exceeds normal variation observed at the population level. This study evaluates NfL and GFAP as diagnostic SRC biomarkers by comparing preseason and post-SRC levels measured up to 28 days after injury to population-based reference intervals. Preseason (uninjured) and post-SRC plasma samples from the Surveillance in High Schools and Community Sports to Reduce Concussions and their Consequences study were quantified with Quanterix's Neurology-4-plex-B assay and compared to previously generated age-matched reference interval. A total of 658 preseason specimens (440 male [67%]; median [range] age, 15.9 [10.4-18.4] years) and 134 post-SRC specimens (72 male [54%]; median [range] age, 16.1 [11.2-18.4] years) were assessed. Median biomarker levels were higher at post-SRC than preseason for both NfL (preseason/post-SRC = 5.1/5.5 pg/ml, P = 0.0339) and GFAP (preseason/post-SRC=51.7/59.7 pg/ml, P < 0.0001). However, when compared to population controls across the same age range, 580 (88%) preseason and 119 (89%) of post-SRC measures fell within the reference interval (5th-95th percentile) for NfL and 644 (98%) preseason and 132 (99%) of post-SRC measures fell within the reference interval for GFAP. Further, the number of participants within or above the reference interval (>95th percentile) did not differ between preseason and post-SRC groups for either NfL (P > 0.9999) or GFAP (P > 0.9999). Body mass index was inversely associated with the distribution of NfL (5th-50th/50th-95th = 23.02/22.15, P = 0.0083; 5th-50th/50th-95th = 24.37/21.74, P = 0.0038) and GFAP (5th-50th/50th-95th = 23.12/22.29, P = 0.0219; 5th-50th/50th-95th = 25.03/22.27, P = 0.0025) within the reference interval, which may explain the skew of NfL and GFAP levels in the SHRed participants compared to population controls at both preseason and post-SRC. We conclude that plasma NfL and GFAP measured up to 28 days post-SRC do not meet criteria for clinical significance as stand-alone post-acute diagnostic biomarkers for adolescent SRC.