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Cited by 209Alzheimer's disease research and treatmentsDementia and Cognitive Impairment ResearchCerebrospinal fluid and hydrocephalus

The DOI matches its Crossref record; 60 of the 60 references it cites were found in a registry or an index.

Characterization of Novel CSF Tau and ptau Biomarkers for Alzheimer’s Disease

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Meredith Jr., Jere E., Sankaranarayanan, Sethu, Guss, Valerie, Lanzetti, Anthony J., Berisha, Flora, Neely, Robert J., Slemmon, J. Randall, Portelius, Erik, Zetterberg, Henrik, Blennow, Kaj, Soares, Holly, Ahlijanian, Michael, Albright, Charles F.·2013·PLoS ONE
DOI 10.1371/journal.pone.0076523PMC PMC3792042
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Abstract

Cerebral spinal fluid (CSF) Aβ42, tau and p181tau are widely accepted biomarkers of Alzheimer's disease (AD). Numerous studies show that CSF tau and p181tau levels are elevated in mild-to-moderate AD compared to age-matched controls. In addition, these increases might predict preclinical AD in cognitively normal elderly. Despite their importance as biomarkers, the molecular nature of CSF tau and ptau is not known. In the current study, reverse-phase high performance liquid chromatography was used to enrich and concentrate tau prior to western-blot analysis. Multiple N-terminal and mid-domain fragments of tau were detected in pooled CSF with apparent sizes ranging from <20 kDa to ~40 kDa. The pattern of tau fragments in AD and control samples were similar. In contrast, full-length tau and C-terminal-containing fragments were not detected. To quantify levels, five tau ELISAs and three ptau ELISAs were developed to detect different overlapping regions of the protein. The discriminatory potential of each assay was determined using 20 AD and 20 age-matched control CSF samples. Of the tau ELISAs, the two assays specific for tau containing N-terminal sequences, amino acids 9-198 (numbering based on tau 441) and 9-163, exhibited the most significant differences between AD and control samples. In contrast, CSF tau was not detected with an ELISA specific for a more C-terminal region (amino acids 159-335). Significant discrimination was also observed with ptau assays measuring amino acids 159-p181 and 159-p231. Interestingly, the discriminatory potential of p181 was reduced when measured in the context of tau species containing amino acids 9-p181. Taken together, these results demonstrate that tau in CSF occurs as a series of fragments and that discrimination of AD from control is dependent on the subset of tau species measured. These assays provide novel tools to investigate CSF tau and ptau as biomarkers for other neurodegenerative diseases.

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Figure 1 Detection of tau fragments in human CSF. Human control and AD CSF subjected to RP-HPLC, fractions collected and run on SDS-PAGE gels followed by western-blotting with different tau antibodies. A) HT7 (mid domain antibody). B) Tau 12 (N-terminal antibody). C) K9JA (C-terminal microtubule repeat domain antibody). D) IgG1 isotype control. On each blot, human recombinant tau441 (tau) is included in lane 1 and molecular weight markers (mw) in lane 2 followed by the HPLC fractions from 1 to 6 or HPLC fractions 7 to 11. Fractions 1 and 2 were pooled and run as a single sample, while fractions 3-10 were run as individual samples. Control CSF (C) and AD CSF (D) samples for each fraction were run side by side for comparison.
Figure 1 Detection of tau fragments in human CSF. Human control and AD CSF subjected to RP-HPLC, fractions collected and run on SDS-PAGE gels followed by western-blotting with different tau antibodies. A) HT7 (mid domain antibody). B) Tau 12 (N-terminal antibody). C) K9JA (C-terminal microtubule repeat domain antibody). D) IgG1 isotype control. On each blot, human recombinant tau441 (tau) is included in lane 1 and molecular weight markers (mw) in lane 2 followed by the HPLC fractions from 1 to 6 or HPLC fractions 7 to 11. Fractions 1 and 2 were pooled and run as a single sample, while fractions 3-10 were run as individual samples. Control CSF (C) and AD CSF (D) samples for each fraction were run side by side for comparison.CC BY · PubMed Central
Figure 2 Tau and ptau ELISAs. Schematic of tau 441 protein with the approximate location of various linear epitope antibodies Tau12, HT7, BT2, Tau5, AT120 and 77G7 and phospho-site specific antibodies AT270 (p181) and PHF6 (p231) indicated; additional antibody epitope and clone information included in Table 1. Antibody combinations used for the different tau and ptau ELISAs are shown. For each assay, capture antibodies are highlighted in red, detection antibodies in black and the minimal tau region required (aa numbering based on tau 441) is indicated. Antibodies used in the INNOTEST/INNO-BIA AlzBio3 total tau and p181 tau assays are also shown for comparison.
Figure 2 Tau and ptau ELISAs. Schematic of tau 441 protein with the approximate location of various linear epitope antibodies Tau12, HT7, BT2, Tau5, AT120 and 77G7 and phospho-site specific antibodies AT270 (p181) and PHF6 (p231) indicated; additional antibody epitope and clone information included in Table 1. Antibody combinations used for the different tau and ptau ELISAs are shown. For each assay, capture antibodies are highlighted in red, detection antibodies in black and the minimal tau region required (aa numbering based on tau 441) is indicated. Antibodies used in the INNOTEST/INNO-BIA AlzBio3 total tau and p181 tau assays are also shown for comparison.CC BY · PubMed Central
Figure 3 Characterization of tau ELISAs. Representative tau 441 standard curves (left panels) and CSF dilution linearity results (right panels) shown for A) HT7-BT2, B) HT7-Tau5, C) Tau12-BT2 and D) Tau12-HT7 tau ELISAs. On each standard curve graph, tau 441 calibrators (Standards) and results for CSF sample dilutions (Samples) are shown. The assay lower limit of quantitation (LLQ, vertical dashed line) is also indicated. On each dilution linearity graph, dilution-corrected tau levels for a pooled control CSF sample and a pooled AD CSF sample relative to sample dilution are shown. The vertical dashed lines indicate the dilution determined to be optimal for CSF analysis.
Figure 3 Characterization of tau ELISAs. Representative tau 441 standard curves (left panels) and CSF dilution linearity results (right panels) shown for A) HT7-BT2, B) HT7-Tau5, C) Tau12-BT2 and D) Tau12-HT7 tau ELISAs. On each standard curve graph, tau 441 calibrators (Standards) and results for CSF sample dilutions (Samples) are shown. The assay lower limit of quantitation (LLQ, vertical dashed line) is also indicated. On each dilution linearity graph, dilution-corrected tau levels for a pooled control CSF sample and a pooled AD CSF sample relative to sample dilution are shown. The vertical dashed lines indicate the dilution determined to be optimal for CSF analysis.CC BY · PubMed Central
Figure 4 Characterization of HT7+77G7 tau ELISA. Representative tau 441 standard curve (left panel) and CSF dilution linearity results (right panel) shown. On the standard curve graph, tau 441 calibrators (Standards) and results for CSF sample dilutions (Samples) are shown. The assay lower limit of quantitation (LLQ, vertical dashed line) is also indicated. On the dilution linearity graph, tau levels for a pooled control and pooled AD CSF samples tested with or without a 100 pg/ml tau 441 spike are shown.
Figure 4 Characterization of HT7+77G7 tau ELISA. Representative tau 441 standard curve (left panel) and CSF dilution linearity results (right panel) shown. On the standard curve graph, tau 441 calibrators (Standards) and results for CSF sample dilutions (Samples) are shown. The assay lower limit of quantitation (LLQ, vertical dashed line) is also indicated. On the dilution linearity graph, tau levels for a pooled control and pooled AD CSF samples tested with or without a 100 pg/ml tau 441 spike are shown.CC BY · PubMed Central
Figure 5 Characterization of ptau assays. Representative ptau standard curves (left panels) and CSF dilution linearity results (right panels) shown for A) HT7-AT270, B) HT7-PHF6, and C) Tau12-AT270 ptau ELISAs. On each standard curve graph, ptau calibrators (Standards) and results for CSF sample dilutions (Samples) are shown. The assay lower limit of quantitation (LLQ, vertical dashed line) is also indicated. On each dilution linearity graph, dilution-corrected ptau levels for a pooled control and a pooled AD CSF samples relative to sample dilution are shown. The vertical dashed lines indicate the dilution determined to be optimal for CSF analysis.
Figure 5 Characterization of ptau assays. Representative ptau standard curves (left panels) and CSF dilution linearity results (right panels) shown for A) HT7-AT270, B) HT7-PHF6, and C) Tau12-AT270 ptau ELISAs. On each standard curve graph, ptau calibrators (Standards) and results for CSF sample dilutions (Samples) are shown. The assay lower limit of quantitation (LLQ, vertical dashed line) is also indicated. On each dilution linearity graph, dilution-corrected ptau levels for a pooled control and a pooled AD CSF samples relative to sample dilution are shown. The vertical dashed lines indicate the dilution determined to be optimal for CSF analysis.CC BY · PubMed Central
Figure 6 Tau and ptau levels in 20 AD and 20 control CSF samples. A set of 20 AD and 20 age-matched normal control CSF samples were analyzed using the tau ELISAs (HT7-BT2, HT7-Tau5, Tau12-BT2, Tau12-HT7 and HT7-77G7) and pTau ELISAs (HT7-AT270, HT7-PHF6 and Tau12-AT270). Dashed lines indicate the assay LLQ corrected for CSF dilution. Statistics based on 2-tailed Student’s t test comparison of log-transformed data, * p < 0.05; ** p< 0.01; *** p < 0.001.
Figure 6 Tau and ptau levels in 20 AD and 20 control CSF samples. A set of 20 AD and 20 age-matched normal control CSF samples were analyzed using the tau ELISAs (HT7-BT2, HT7-Tau5, Tau12-BT2, Tau12-HT7 and HT7-77G7) and pTau ELISAs (HT7-AT270, HT7-PHF6 and Tau12-AT270). Dashed lines indicate the assay LLQ corrected for CSF dilution. Statistics based on 2-tailed Student’s t test comparison of log-transformed data, * p < 0.05; ** p< 0.01; *** p < 0.001.CC BY · PubMed Central

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Metadata

Authors
Meredith Jr., J. E., Sankaranarayanan, S., Guss, V., Lanzetti, A. J., Berisha, F., Neely, R. J., Slemmon, J. R., Portelius, E., Zetterberg, H., Blennow, K., Soares, H., Ahlijanian, M., & Albright, C. F.
Affiliations
  • Bristol-Myers Squibb (United States) (US)
  • University of Gothenburg (SE)
  • UCL Queen Square Institute of Neurology (GB)
ORCID
Venue
PLoS ONE
Publisher
Public Library of Science (PLoS)
Published
2013-10-07
Volume
8
Issue
10
Pages
e76523
ISSN
1932-6203 (ISSN-L)
DOI
10.1371/journal.pone.0076523
PMC
PMC3792042
OpenAlex
W2024790026
Cited by
209
References
60
Citations by year
2026: 6 · 2025: 13 · 2024: 14 · 2023: 14 · 2022: 11 · 2021: 19 · 2020: 23 · 2019: 28 · 2018: 29 · 2017: 16 · 2016: 11 · 2015: 13 · 2014: 11
FWCI
8.10
Type
journal article
Language
en
Open access
Licence
Locations
  • PLoS ONE (publishedVersion, cc-by)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • DOAJ (DOAJ: Directory of Open Access Journals) (submittedVersion, cc-by-sa)
  • UCL Discovery (University College London) (submittedVersion)
  • Figshare (submittedVersion, cc-by)
Funders
  • Bristol-Myers Squibb
  • Vetenskapsrådet
Topics
  • Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Alzheimer's disease research and treatments
  • Health Sciences › Medicine › Psychiatry and Mental health › Dementia and Cognitive Impairment Research
  • Health Sciences › Medicine › Clinical Neurology › Cerebrospinal fluid and hydrocephalus
Keywords
Alzheimer's disease, cerebrospinal fluid, tau protein, phosphorylated tau, biomarkers, preclinical Alzheimer's disease, tau isoforms, diagnostic discrimination
MeSH terms
Aged, Aged, 80 and over, Alzheimer Disease, Chromatography, High Pressure Liquid, Enzyme-Linked Immunosorbent Assay, Humans, Male, Middle Aged, Molecular Weight, Peptide Fragments, Phosphorylation, Sensitivity and Specificity, Blotting, Western, Reproducibility of Results, Biomarkers, tau Proteins, Chromatography, Reverse-Phase

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