Simonsen, Anja H5; Bahl, Justyna M C4; Danborg, Pia B4; Lindstrom, Veronica4; Larsen, Severin O4; Grubb, Anders4; Heegaard, Niels Henrik Helweg6; Waldemar, Gunhild7
1 Section of Teaching, Department of Biomedical Sciences, Faculty of Health and Medical Sciences, Københavns Universitet2 Department of Clinical Medicine, Department of Clinical Medicine, Faculty of Health and Medical Sciences, Københavns Universitet3 Department of Clinical Medicine, Faculty of Health and Medical Sciences, Københavns Universitet4 unknown5 Department of Clinical Medicine, Faculty of Health and Medical Sciences, Københavns Universitet6 Section of Teaching, Department of Biomedical Sciences, Faculty of Health and Medical Sciences, Københavns Universitet7 Department of Clinical Medicine, Department of Clinical Medicine, Faculty of Health and Medical Sciences, Københavns Universitet
Cerebrospinal fluid (CSF) is a potential source for new biomarkers due to its proximity to the brain. This study aimed to clarify the stability of the CSF proteome when undergoing pre-analytical factors. We investigated the effects of repeated freeze/thaw cycles, protease inhibitors and delayed storage for 4h, 24h or 14 days at -20°C, 4°C and room temperature (RT) after centrifugation compared with our standard practice of two hours at RT before placing the samples in an -80°C environment. The results were obtained using immunoassays for amyloid-beta 1-42 (Aβ42), tau, phosphorylated tau (P-tau) and cystatin C and using surface-enhanced laser desorption/ionisation time-of-flight (SELDI-TOF) mass spectrometry for proteomic profiling. Tau and P-tau were susceptible to repeated freeze/thaw cycles while SELDI-TOF analysis produced eight significant peaks and additional artefact peaks from samples with added protease inhibitors. Delayed storage for different durations and in different temperatures produced six significant SELDI-TOF peaks. Aβ42 and tau were susceptible to increased temperatures and the duration before storage, whereas P-tau and cystatin C were not. Transthyretin and several of its isoforms were found using SELDI-TOF and were susceptible to freeze/thaw cycles and to increased temperature and length of time prior to storage. We recommend that CSF should be collected and centrifuged immediately after sampling and prior to storage at -80°C without the addition of protease inhibitors. Freeze/thawing should be avoided because of the instability of tau, P-tau and transthyretin. Standardised CSF sampling, handling and storage for biomarker research are essential for accurately comparing the results obtained by different studies and institutions.
Journal of Neuroscience Methods, 2013, Vol 215, Issue 2, p. 234-240