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Mol Neurobiol. 2016 May;53(4):2189-99. doi: 10.1007/s12035-015-9167-5. Epub 2015 May 7.

Validation of 14-3-3 Protein as a Marker in Sporadic Creutzfeldt-Jakob Disease Diagnostic.

Author information

1
Department of Neurology, University Medical Center Göttingen and German Center for Neurodegenerative Diseases (DZNE)-site Göttingen, Robert-Koch Str. 40, 37075, Göttingen, Germany. Matthias.Schmitz@med.uni-goettingen.de.
2
Department of Neurology, University Medical Center Göttingen and German Center for Neurodegenerative Diseases (DZNE)-site Göttingen, Robert-Koch Str. 40, 37075, Göttingen, Germany.
3
Department of Pathology, The University of Melbourne, Parkville, 3010, Australia.
4
Chronic Disease Programme, CIBERNED, and CIEN Foundation-Queen Sofia Foundation, Instituto de Salud Carlos III, Madrid, Spain.
5
Pharmaceutical Sciences, Laboratory of Pharmacology, Aristotle University of Thessaloniki, Thessaloniki, 54124, Greece.
6
Service de Biochimie et Biologie Moléculaire, Hôpital Lariboisière, Assistance Publique - Hôpitaux de Paris, 2, rue A Paré, 75475, Paris cedex 10, France.
7
Department of Molecular Pathology and Neuropathology, Medical University of Lodz, 8/10 Czechoslowacka Str., 92-216, Lodz, Poland.
8
Center for Neuroscience and Cell Biology, Faculty of Medicine, University of Coimbra, Coimbra, Portugal.
9
Department of Molecular Microbiology and Immunology, Nagasaki University Graduate School of Biomedical Sciences, Sakamoto 1-12-4, Nagasaki, Japan.
10
Neurology Department, Hospital Clinic, IDIBAPS, Barcelona, Spain.
11
Department of Cell Biology and Neurosciences, Istituto Superiore di Sanità, Rome, Italy.
12
Department of Multidisiplinary Laboratory Medicine and Medical Biochemistry Division of Diagnostics and Technology Akerhus University Hospital 1478 Lørenskog, 1478, Lørenskog, Norway.
13
The Public Health Agency of Sweden, Unit for highly pathogenic viruses, 171 82, Solna, Sweden.
14
Department of Prion Diseases, Slovak Medical University Bratislava, Limbová 14, 833-03, Bratislava, Slovakia.
15
Ilsong Institute of Life Science, College of Medicine, Hallym University, Anyang, Republic of Korea.
16
Western General Hospital, National Creutzfeldt-Jakob Disease Surveillance Unit, Crewe Road, Edinburgh, EH4 2XU, UK.

Abstract

At present, the testing of 14-3-3 protein in cerebrospinal fluid (CSF) is a standard biomarker test in suspected sporadic Creutzfeldt-Jakob disease (sCJD) diagnosis. Increasing 14-3-3 test referrals in CJD reference laboratories in the last years have led to an urgent need to improve established 14-3-3 test methods. The main result of our study was the validation of a commercially available 14-3-3 ELISA next to the commonly used Western blot method as a high-throughput screening test. Hereby, 14-3-3 protein expression was quantitatively analyzed in CSF of 231 sCJD and 2035 control patients. We obtained excellent sensitivity/specificity values of 88 and 96% that are comparable to the established Western blot method. Since standard protocols and preanalytical sample handling have become more important in routine diagnostic, we investigated in a further step the reproducibility and stability of 14-3-3 as a biomarker for human prion diseases. Ring trial data from 2009 to 2013 revealed an increase of Fleiss' kappa from 0.51 to 0.68 indicating an improving reliability of 14-3-3 protein detection. The stability of 14-3-3 protein under short-term and long-term storage conditions at various temperatures and after repeated freezing/thawing cycles was confirmed. Contamination of CSF samples with blood appears likely to be an important factor at a concentration of more than 2500 erythrocytes/μL. Hemolysis of erythrocytes with significant release of 14-3-3 protein started after 2 days at room temperature. We first define clear standards for the sample handling, short- and long-term storage of CSF samples as well as the handling of blood- contaminated samples which may result in artificially elevated CSF levels of 14-3-3.

KEYWORDS:

14-3-3 protein; Biomarker; Cerebrospinal fluid; Creutzfeldt-Jakob disease; Pre-mortem test; Standardization

PMID:
25947081
DOI:
10.1007/s12035-015-9167-5
[Indexed for MEDLINE]

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