RACE Study: Rapid, Accurate and Cost-effective Analysis of Glial Fibrillary Acid Protein Using a Hand-held Biosensor for Patient With Concussion in Acute Care and at Home Monitoring

BACKGROUND

It is estimated 100-300/100,000 people worldwide present to a hospital with traumatic brain injury (TBI) annually, the majority of which are classified as mild. TBI is a disruption in normal brain function caused by external biomechanical forces transmitted directly or indirectly to the head, and is a leading cause of death and disability in Canada. Approximately 1 in 450 Canadians report brain injury as their most significant injury associated with disability in the previous year. Mild traumatic brain injury (mTBI) is operationalized under clinical severity by a Glasgow Coma Scale (GCS) score of 13-15 and is often used interchangeably with concussion, though the most recent consensus definition of concussion is precluded by positive findings on standard neuroimaging techniques. Although labelled as mild with typical recovery times within two weeks of injury for adults and four weeks for youth, up to 30% of patients with concussion experience prolonged symptoms (headache, fatigue, dizziness, insomnia, depression, anxiety, poor balance, and cognitive deficits, etc.) contributing to significant functional impairment and disease burden. Furthermore, in 2016 approximately 10% of diagnosed concussions in Ontario returned to the emergency department within 14 days of injury, increasing demands on the health care system.

Lending to its identity as one of the most complex injuries to diagnose and manage, concussion currently relies on subjective measures and symptom reports as clinical indices of injury. There has been accelerated interest in addressing this limitation through research efforts working to establish objective measures of injury including advanced neuroimaging imaging techniques, machine learning of basic physiological functions (brain waves, heart rate, blood pressure, etc.), and blood biomarkers. Blood biomarkers have shown promising results regarding their ability to detect or predict severe and moderate TBI, but results in mTBI are mixed and require further investigation. Two biomarkers of brain injury - glial fibrillary acidic protein (GFAP) and ubiquitin c-terminal hydrolase L1 (UCH-L1) - were recently FDA approved to help identify necessity of a CT scan for adult mTBI patients who might have intracranial lesions. However, beyond the currently insufficient level of evidence regarding blood biomarker applications for concussion diagnosis or prognosis, an additional hurdle for future implementation in a clinical setting remains the high cost and time-consuming assay methodologies for marker detection. Single molecule array technology (SIMOA) is a fully automated immunoassay capable of biomarker detection at the femtogram level, approximately 900x more sensitive than conventional enzyme-linked immunosorbent assays. As the current gold-standard for low concentration biomarker detection, significant monetary and procedural costs may limit SIMOA's future applications for concussion biomarker detection in clinical settings. Fortunately, technological advancements are pushing the boundaries of conventional assay approaches to minimize cost and maximize efficiency, progressing towards point-of-care assay devices.

The investigators have developed a GFAP nano-biosensor capable of measuring GFAP concentrations in similar orders of magnitude as SIMOA. Briefly, the GFAP electrochemical biosensor was developed using nano-porous carbon on screen-printed electrodes with hydroxylamine (NH2OH). The nano-coated and functionalized electrodes were immobilized with monoclonal anti-human GFAP capture antibody, then blocked with bovine serum albumin. GFAP binding frequency was translated to serum concentration levels using electrochemical impedance spectroscopy (EIS). The nano-porous biosensor provided ultrasensitive GFAP detection in a wide operational range of 100 fg/mL - 10 ng/mL concentrations in the human serum. It selectively detected GFAP when tested against other biomarkers released after brain injury, and revealed a clinical sensitivity for detection of patients across TBI severities (total n=70; TBI n=26, healthy control n=44) at 84.62% (95%CI, 65.1% to 95.6%) and specificity at 61.36% (CI, 45.5% to 75.6%). The limit of detection of the GFAP biosensor was measured to be 86.6 fg/mL in serum. Additionally, our nano-biosensor demonstrated <2% intra-variation and <1% inter-variation, tackling the concern of reproducible biosensor production needed for clinical detection. This research seeks to validate the nano-biosensor in concussion patients presenting to the Emergency Department.

OBJECTIVES