High sensitivity of an ELISA kit for detection of the gamma-isoform of 14-3-3 proteins: usefulness in laboratory diagnosis of human prion disease

Yuki Matsui,Kazuo Mutsukura,Katsuya Satoh,Susumu Shirabe,Toshiaki Miyazaki,Ryuichiro Atarashi,Akira Satoh,Yasufumi Kataoka,Noriyuki Nishida

doi:10.1186/1471-2377-11-120

Yuki Matsui, Kazuo Mutsukura + Show 7 more

Open Access

https://doi.org/10.1186/1471-2377-11-120

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Abstract

BackgroundThe gamma-isoform of the 14-3-3 protein (14-3-3 gamma) is expressed in neurons, and could be a specific marker for neuronal damage. This protein has been reported as a detectable biomarker, especially in the cerebrospinal fluid (CSF) of Creutzfeldt-Jakob disease (CJD) patients by Western blotting (WB) or enzyme-linked immunosorbent assays (ELISAs). Western blotting for 14-3-3 gamma is not sensitive, and the reported data are conflicting among publications. An ELISA specific for 14-3-3 gamma is not available.MethodsCJD patients (n = 114 sporadic CJD patients, 7 genetic CJD, and 3 iatrogenic CJD) and 99 patients with other neurodegenerative diseases were examined in this study. The CSF samples obtained were analyzed by Western blotting for 14-3-3 gamma, and by ELISA for total tau protein. We evaluated the sensitivity and specificity of the newly developed sandwich ELISA for 14-3-3 gamma.ResultsThe cut-off value of the 14-3-3 gamma ELISA was > 1, 683 AU/ml; and sensitivity was 95.2%, with 72.7% specificity. This specificity was the same for the total tau protein ELISA. Seven CJD cases were negative by WB but positive using the 14-3-3 gamma ELISA, indicating that the ELISA is more sensitive. All 21 cases of early stage CJD could be diagnosed using a combination of the 14-3-3γ ELISA and diffusion weighted MR imaging (DWI-MRI).ConclusionThe 14-3-3 gamma ELISA was more sensitive than conventional WB, and was useful for laboratory diagnosis of CJD, similar to the ELISA for the tau protein. Using DWI-MRI and these ELISA tests on CSF, diagnosis of CJD will be possible even at early stages of the disease.

Highlights

The gamma-isoform of the 14-3-3 protein (14-3-3 gamma) is expressed in neurons, and could be a specific marker for neuronal damage
Detection of total tau protein in cerebrospinal fluid (CSF) Samples The levels of total tau protein in CSF were determined in 223 patients, and significant differences were observed among individuals (Tables 1 and 2)
There were two problems with our study: 20 dementia of Alzheimer’s type (DAT) cases identified that the level of enzyme-linked immunosorbent assays (ELISAs) was 3, 000-5, 000 AU/ml, but they could not be detected by Western blotting (WB) and these results showed a discrepancy between the 14-3-3g WB and ELISA; and the specificity of the ELISA was very low (Figure 1)

Summary

Introduction

The gamma-isoform of the 14-3-3 protein (14-3-3 gamma) is expressed in neurons, and could be a specific marker for neuronal damage. This protein has been reported as a detectable biomarker, especially in the cerebrospinal fluid (CSF) of Creutzfeldt-Jakob disease (CJD) patients by Western blotting (WB) or enzyme-linked immunosorbent assays (ELISAs). Hshich et al.[1] reported use of the 14-3-3 protein for diagnosis of prion diseases in 1996 This protein is a reliable marker of rapid neuronal destruction, and has been detected in the cerebrospinal fluid (CSF) of several progressive neurological disorders. The 14-3-3 protein has been reported to be a detectable biomarker, especially in the CSF of CJD patients, by WB or enzyme-linked immunosorbent assay (ELISA). We attempted to improve upon these four problems and attempted to develop a new, specific, 14-3-3g ELISA

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