title: Sporadic Creutzfeldt-Jakob disease diagnostic accuracy is improved by a new CSF ELISA 14-3-3γ assay.
abstract: Protein 14-3-3 is a reliable marker of rapid neuronal damage, specifically increased in cerebrospinal fluid (CSF) of sporadic Creutzfeldt-Jakob disease (sCJD) patients. Its detection is usually performed by Western Blot (WB), prone to methodological issues. Our aim was to evaluate the diagnostic performance of a recently developed quantitative enzyme-linked immunosorbent (ELISA) assay for 14-3-3γ, in comparison with WB and other neurodegeneration markers. CSF samples from 145 patients with suspicion of prion disease, later classified as definite sCJD (n=72) or Non-prion diseases (Non-CJD; n=73) comprised our population. 14-3-3 protein was determined by WB and ELISA. Total Tau (t-Tau) and phosphorylated Tau (p-Tau) were also evaluated. Apolipoprotein E gene (ApoE) and prionic protein gene (PRNP) genotyping was assessed. ELISA 14-3-3γ levels were significantly increased in sCJD compared to Non-CJD patients (p<0.001), showing very good accuracy (AUC=0.982; sensitivity=97%; specificity=94%), and matching WB results in 81% of all cases. It strongly correlated with t-Tau and p-Tau (p<0.0001), showing slightly higher specificity (14-3-3 WB - 63%; Tau - 90%; p-Tau/t-Tau ratio - 88%). From WB inconclusive results (n=44), ELISA 14-3-3γ correctly classified 41 patients. Additionally, logistic regression analysis selected ELISA 14-3-3γ as the best single predictive marker for sCJD (overall accuracy=93%). ApoE and PRNP genotypes did not influence ELISA 14-3-3γ levels. Despite specificity for 14-3-3γ isoform, ELISA results not only match WB evaluation but also help discrimination of inconclusive results. Our results therefore reinforce this assay as a single screening test, allowing higher sample throughput and unequivocal results.
authors.affiliation: Neurochemistry Laboratory, Neurology Department, University Hospital Coimbra, Centro Hospitalar e Universitário de Coimbra, Praceta Mota Pinto, 3000-075 Coimbra, Portugal; Faculty of Medicine, University of Coimbra, Portugal; CNC-Center for Neuroscience and Cell Biology, University of Coimbra, Rua Larga, Faculty of Medicine, Pólo I, 1st Floor, 3004-504 Coimbra, Portugal. Electronic address: jajao86@gmail.com.
authors.forename: M J
authors.initials: MJ
authors.lastname: Leitão
authors.affiliation: Neurochemistry Laboratory, Neurology Department, University Hospital Coimbra, Centro Hospitalar e Universitário de Coimbra, Praceta Mota Pinto, 3000-075 Coimbra, Portugal; Faculty of Medicine, University of Coimbra, Portugal; CNC-Center for Neuroscience and Cell Biology, University of Coimbra, Rua Larga, Faculty of Medicine, Pólo I, 1st Floor, 3004-504 Coimbra, Portugal.
authors.forename: I
authors.initials: I
authors.lastname: Baldeiras
authors.affiliation: CNC-Center for Neuroscience and Cell Biology, University of Coimbra, Rua Larga, Faculty of Medicine, Pólo I, 1st Floor, 3004-504 Coimbra, Portugal.
authors.forename: M R
authors.initials: MR
authors.lastname: Almeida
authors.affiliation: Neurochemistry Laboratory, Neurology Department, University Hospital Coimbra, Centro Hospitalar e Universitário de Coimbra, Praceta Mota Pinto, 3000-075 Coimbra, Portugal; Faculty of Medicine, University of Coimbra, Portugal.
authors.forename: M H
authors.initials: MH
authors.lastname: Ribeiro
authors.affiliation: CNC-Center for Neuroscience and Cell Biology, University of Coimbra, Rua Larga, Faculty of Medicine, Pólo I, 1st Floor, 3004-504 Coimbra, Portugal.
authors.forename: A C
authors.initials: AC
authors.lastname: Santos
authors.affiliation: CNC-Center for Neuroscience and Cell Biology, University of Coimbra, Rua Larga, Faculty of Medicine, Pólo I, 1st Floor, 3004-504 Coimbra, Portugal.
authors.forename: M
authors.initials: M
authors.lastname: Ribeiro
authors.affiliation: Neurology Department, University Hospital Coimbra, Centro Hospitalar e Universitário de Coimbra, Praceta Mota Pinto, 3000-075 Coimbra, Portugal.
authors.forename: J
authors.initials: J
authors.lastname: Tomás
authors.affiliation: Neurology Department, St Marcos Hospital, Braga, Portugal.
authors.forename: S
authors.initials: S
authors.lastname: Rocha
authors.affiliation: Neurology Department, University Hospital Coimbra, Centro Hospitalar e Universitário de Coimbra, Praceta Mota Pinto, 3000-075 Coimbra, Portugal; Faculty of Medicine, University of Coimbra, Portugal; CNC-Center for Neuroscience and Cell Biology, University of Coimbra, Rua Larga, Faculty of Medicine, Pólo I, 1st Floor, 3004-504 Coimbra, Portugal.
authors.forename: I
authors.initials: I
authors.lastname: Santana
authors.affiliation: Neurochemistry Laboratory, Neurology Department, University Hospital Coimbra, Centro Hospitalar e Universitário de Coimbra, Praceta Mota Pinto, 3000-075 Coimbra, Portugal; Neurology Department, University Hospital Coimbra, Centro Hospitalar e Universitário de Coimbra, Praceta Mota Pinto, 3000-075 Coimbra, Portugal; Faculty of Medicine, University of Coimbra, Portugal; CNC-Center for Neuroscience and Cell Biology, University of Coimbra, Rua Larga, Faculty of Medicine, Pólo I, 1st Floor, 3004-504 Coimbra, Portugal.
authors.forename: C R
authors.initials: CR
authors.lastname: Oliveira
mesh_terms: D048948:14-3-3 Proteins / Q000134:cerebrospinal fluid*; D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D001057:Apolipoproteins E / Q000235:genetics; D015415:Biomarkers / Q000134:cerebrospinal fluid; D015153:Blotting, Western; D002648:Child; D007562:Creutzfeldt-Jakob Syndrome / Q000134:cerebrospinal fluid* / Q000235:genetics; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D010766:Phosphorylation; D000072002:Prion Proteins / Q000235:genetics; D012680:Sensitivity and Specificity; D055815:Young Adult; D016875:tau Proteins / Q000134:cerebrospinal fluid
chemical_list: D048948:14-3-3 Proteins; D001057:Apolipoproteins E; D015415:Biomarkers; C054369:MAPT protein, human; C512656:PRNP protein, human; D000072002:Prion Proteins; D016875:tau Proteins
keywords: 14-3-3 protein; CSF; ELISA; Tau proteins; Western Blot; sCJD