This paper highlights the problems associated with quantifying the levels of serum antibodies recombinant glutathione S-transferase (GST). Measurement of anti-GST antibody in traditional immunoassays, where GST is bound directly to the ELISA plate, was found to underestimate the amount of specific antibody levels in test sera-GST. This insensitivity of immunoassay Anti-GST Antibody can be overcome by using one of several GST fusion recombinant proteins as antigen coating ELISA rather than simply GST. Comparison of anti-GST securities valued by the two procedures indicated that the use of underestimating unfused GST as the anti-GST antibody by more than ten times.
Creutzfeldt-Jakob disease (CJD) is a rapidly progressing disease and fatal central nervous system appears to be caused by the prion. The clinical syndrome of CJD include a progressive dementia fast, myoclonus, visual or cerebellar, pyramidal or extrapyramidal signs and akinetic mutism. To date, a definitive diagnosis can be made by neuropathological examination and demonstration of the pathological isoform of the prion protein (PrPSc) in the central nervous tissue or a biopsy or autopsy. In patients with new variant CJD (vCJD), a pathological isoform was also detected in the tonsils. Probable or possible diagnosis of CJD, especially sporadic CJD (CJD) can be achieved according intravitally clinical manifestation, typical EEG changes and the appearance or the alternation of some neuronal proteins in cerebrospinal fluid ( CSF). However, only for 14-3-3 CSF immunoblot is included in the diagnostic criteria despite the fact that other surrogate markers may also have a high potential for differential diagnosis.
Tau is a microtubule-associated protein capable of promoting microtubule assembly and stability in the nervous system. In the brains of normal adult humans, six isoforms of tau have been reported, which are encoded by a single gene with 16 exons by alternative mRNA splicing. They differ from each other by the presence or absence of an insertion of 29 amino acids (encoded by exon-2) or the insertion of 58 amino acids (encoded by exon-2 and – 3) in the amino-terminal half of the protein and the presence or absence of a repeated amino acid 31 (encoded by exon-10) in its carboxy-terminal half. Based on the constitution of the exon-2, -3 and -10, tau, tau mature proteins have different lengths, including Tau352, Tau381, Tau383, Tau410, and Tau412 Tau441. All tau isoforms containing three repeats of other municipalities in its C-terminal half, which are responsible for interaction with microtubules. Therefore, according to the repetition of exon-10 in C-terminal peptide, tau protein can be divided into two groups, 3R and 4R-tau-tau. Based on the presence of the insertion (s) from exon-2 and / or exon-3 in N-terminal protein tau has three forms, namely 1N-tau, tau-0N and 2N-tau. The presence of tau isoforms in the human brain may vary with age, ie the amount of 3R and 4R-tau-tau are similar in the adult cerebral cortex, while 0N-tau is predominant in the brain developing.
Under certain pathological conditions, tau has become the main component of intracellular filamentous deposits, such as Alzheimer’s disease (AD). Proportions of Altered tau isoforms were observed in frontotemporal dementia, Parkinson’s disease linked to chromosome 17 (FTDP-17) and Pick’s disease. Promising results of the diagnostic sensitivity and specificity of tau-protein by ELISA in CSF have been reported, which was accepted as a standard procedure in the diagnosis and evaluation of AD therapy. The diagnostic value of CSF tau CJD has been extensively evaluated, showing a clear way for diseases. However, the profiles of the isoforms of tau in the CSF of CJD patients is unknown, possibly due to lack of tau isoforms or exon-specific antibodies.
In this study, human tau exon-2, -3 and -10 fusion proteins and various specific tau isoform were individually expressed in E. coli, using tau exon-2, -3 and -10 fusion proteins as antigens, tau exon-specific polyclonal antibodies were prepared. We confirmed that the GST Antibody prepared to recognize and distinguish tau isoforms containing the various modules of the exon. Based on polyclonal antibodies prepared profiles of tau in CSF samples from 96 Chinese patients with probable sporadic CJD, genetic CJD 6 and 22 cases were excluded from the possibility of CJD were analyzed by Western blot. A 65 KD tau specific band, which was recognized mainly by antibodies against tau Exon-2 and-10, appeared especially in much of the CSF samples of probable cases of sporadic CJD. Our study provides reliable human tau exon specific polyclonal antibodies and 65 KD tau specific band can be used as a new biomarker for the detection of sporadic CJD.
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