Abstracts

Rationale: The principal goals of the STOP SUDEP Program are 1) SUDEP registry, 2) tissue biorepository, and 3) immediate sample utilization for SUDEP research. The scope of molecular investigations and validity of results is crucially dependent on the quality of the available tissue. Multiple factors influence sample diagnostic utility and broadly informed qualitative assessment of the specimen critically dictates investigational and diagnostic value of a specific sample.Methods: STOP SUDEP Program is a national and international collaborative network of families of patients with epilepsy, forensic pathologists, organizations supporting epilepsy research and education, and investigators dedicated to SUDEP research. This Biorepository houses biological specimens that vary in source, conservation methods, collection age, and amount. We performed comprehensive assessment of biospecimens with regards to their integrity, quantity, and quality using traditional analytical methods in parallel with the visual automated fluorescence electrophoresis (VAFE).Results: We extracted DNA from series of archived tissues and performed the first simultaneous quantitative and qualitative analyses of the genomic DNA on the visual automated fluorescence electrophoresis (VAFE), before and following whole genome amplification (WGA), in parallel with traditional quality control (QC) methods. The sample detection limit was >1 ng/mL by the Nanodrop1000 and >0.5 ng/mL by VAFE. However, there was a statistically significant difference in the total DNA per standard sample amount as measured by individual platforms (40 to 384 ng per the Nanodrop1000 versus 12 to 49 ng per the Agilent 2200 TapeStation; t=3.68, P = 0.002). VAFE also clearly discriminated small but individually variable degradation in all sample types, as evidenced by the presence of lower-molecular-weight DNA fragments. Moreover, VAFE QC data correlated with subsequent sample performance in PCR, sequencing, and a high-density comparative genome hybridization array. Conclusions: Conclusions: STOP SUDEP Program is the first SUDEP registry and tissue repository dedicated to concurrent analysis and research of mechanisms and molecular risk factors of SUDEP where pre-analytical tissue quality assessment is built in the evaluation of the clinical utility of samples. Our comparative assessment of SUDEP samples establishes that VAFE is cost effective and informative in the qualitative and quantitative stratification of tissue derived gDNA for the purpose of investigational and diagnostic applications, and it adds to the confidence in the validity of the resultant genetic data. This works was supported by National Institute of Neurological Disorders and Stroke grants NS067013 and NS067013S (A.M.G.), NS-049130, and NS076916 (J.L.N.); CURE; Fiorito Foundation and the Emma Bursick Memorial Fund Q2 (A.M.G.); the Blue Bird Circle Foundation (J.L.N.); and the Stockholm County Council, ALF (T.T., O.S.) Q3. Agilent Technologies loaned the Agilent 2200 TapeStation and gave access to the prototype Agilent Genomic DNA ScreenTape.