High Throughput ELISAs to Measure a Unique Glycan on Transferrin in Cerebrospinal Fluid: A Possible Extension toward Alzheimer's Disease Biomarker Development.

Department of Biochemistry, Fukushima Medical University School of Medicine, 1 Hikarigaoka, Fukushima 960-1295, Japan.
International journal of Alzheimer's disease 01/2011; 2011:352787. DOI: 10.4061/2011/352787
Source: PubMed

ABSTRACT We have established high-throughput lectin-antibody ELISAs to measure different glycans on transferrin (Tf) in cerebrospinal fluid (CSF) using lectins and an anti-transferrin antibody (TfAb). Lectin blot and precipitation analysis of CSF revealed that PVL (Psathyrella velutina lectin) bound an unique N-acetylglucosamine-terminated N-glycans on "CSF-type" Tf whereas SSA (Sambucus sieboldiana agglutinin) bound α2,6-N-acetylneuraminic acid-terminated N-glycans on "serum-type" Tf. PVL-TfAb ELISA of 0.5 μL CSF samples detected "CSF-type" Tf but not "serum-type" Tf whereas SSA-TfAb ELISA detected "serum-type" Tf but not "CSF-type" Tf, demonstrating the specificity of the lectin-TfAb ELISAs. In idiopathic normal pressure hydrocephalus (iNPH), a senile dementia associated with ventriculomegaly, amounts of the SSA-reactive Tf were significantly higher than in non-iNPH patients, indicating that Tf glycan analysis by the high-throughput lectin-TfAb ELISAs could become practical diagnostic tools for iNPH. The lectin-antibody ELISAs of CSF proteins might be useful for diagnosis of the other neurological diseases.

  • [Show abstract] [Hide abstract]
    ABSTRACT: We developed a high-throughput ELISA for measuring α2,6-sialylated transferrin (Tf), based on inhibition of anti-transferrin antibody binding to α2,6-sialylated Tf by a lectin, Sambucus sieboldiana Agglutinin (SSA). The inhibition was not observed with other glycoforms such as periodate-treated, sialidase-treated, and sialidase/galactosidase-treated Tf, suggesting that the assay was glycoform specific. This finding was applied to an automated latex-agglutination immunoassay, using SSA lectin as an inhibitor (SSA-ALI). The concentration of α2,6-sialylated Tf measured by SSA-ALI in human cerebrospinal fluid was correlated with that of ELISA (r(2)=0.8554), previously developed for measuring α2,6-sialylated Tf.
    Journal of Biochemistry 08/2013; · 3.07 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Glycosylation is one of the most common, and the most complex, forms of post-translational modification of proteins. This review serves to highlight the role of protein glycosylation in Alzheimer disease (AD), a topic that has not been thoroughly investigated, although glycosylation defects have been observed in AD patients. The major pathological hallmarks in AD are neurofibrillary tangles and amyloid plaques. Neurofibrillary tangles are composed of phosphorylated tau, and the plaques are composed of amyloid β-peptide (Aβ), which is generated from amyloid precursor protein (APP). Defects in glycosylation of APP, tau and other proteins have been reported in AD. Another interesting observation is that the two proteases required for the generation of amyloid β-peptide (Aβ), i.e. γ-secretase and β-secretase, also have roles in protein glycosylation. For instance, γ-secretase and β-secretase affect the extent of complex N-glycosylation and sialylation of APP, respectively. These processes may be important in AD pathogenesis, as proper intracellular sorting, processing and export of APP are affected by how it is glycosylated. Furthermore, lack of one of the key components of γ-secretase, presenilin, leads to defective glycosylation of many additional proteins that are related to AD pathogenesis and/or neuronal function, including nicastrin, reelin, butyrylcholinesterase, cholinesterase, neural cell adhesion molecule, v-ATPase, and tyrosine-related kinase B. Improved understanding of the effects of AD on protein glycosylation, and vice versa, may therefore be important for improving the diagnosis and treatment of AD patients.
    FEBS Journal 10/2013; · 4.25 Impact Factor


1 Download
Available from