BIOMARKER STABILITY IN CSF: PRE-ANALYTICAL FACTORS IN A PROSPECTIVE COLLECTION

Evaluation of cerebrospinal fluid (CSF) biomarkers in Alzheimer's disease (AD) is becoming increasingly important to improve the reliability of ante-mortem disease diagnosis to ensure proper patient management. Development of highly precise assays for amyloid 1-42 (Aβ42) and tau protein has allowed investigators to better characterize pre-analytical sample handling factors that may affect clinical interpretation. A prospective collection study was designed to model different CSF handling scenarios for storage conditions at the clinical site, shipping, and storage and handling at the testing site. CSF was prospectively collected from 46 healthy individuals under an IRB approved protocol. A 10mL CSF aliquot was collected by lumbar puncture in the L3/L4 or L4/L5 interstitial space. One milliliter aliquots were stored and shipped at: -80°C, -20°C, 4°C and 25°C. This study investigated storage, shipping and handling conditions, including freeze/thaws for 6 randomly selected patients using an immunometic chemiluminescent assay. Additional studies examined the influence of storage tube material. Compared to -80°C, aliquots stored at -20°C and 25°C had Aβ42 losses from 5 to 20% and 40% loss for aliquots stored at ambient temperatures for 48 hours. There was little to no effect on tau concentrations across the range of storage temperatures. Aliquots initially stored at -80°C or ambient temperature showed the greatest decreases in measured Aβ42 levels (10 to 15%) following a second freeze/thaw cycle, whereas no loss of tau was observed with a freeze/thaw cycle. Thawing CSF samples at ambient temperature versus a water bath did not affect concentrations of either biomarker. Results were consistent between patients. Storing samples for 2h at ambient temperature post thaw resulted in less than 10% loss for either marker. The Eppendorf Lobind® tubes allowed transfer of CSF and storage at 4°C for up to six days without significant loss of either Aβ42 or tau biomarkers. Storage and transfer conditions of CSF can have a significant effect on the level of Aβ42 measured but do not impact tau concentrations appreciably. The reliability and utility of these markers in routine testing can be improved by minimizing analyte losses by choice of conditions from collection to testing site.