This antibody was raised against a synthetic peptide corresponding to the amino-acid sequence encoded by exon 10 of Tau proteins. This specific sequence is found in three of the six human brain Tau isoforms (E10+) and its expression is developmentally regulated [1].
Exon 10 expression leads to the presence of a fourth repeat (R2) that increases the affinity of tau for tubulin dimers [2].
Mutations in the intron regulating the splicing of exon 10 or in exon 10 are responsible for diseases named "frontotemporal dementia and Parkinsonism linked to chromosome 17" (FTDP-17) (Spillantini MG, Goedert M, Tau protein pathology in neurodegenerative diseases.Trends Neurosci 1998 Oct;21(10):428-33)
The neat serum is conserved in 50% (v/v) glycerol. It does not contain any other additive or conservative products.
A New Zealand rabbit was immunized sub-cutaneously with a KLH coupled synthetic peptide (Neosystems, France).
This antibody can be used for western blotting. It is difficult to use at the immunohistochemical level.
1. Western-blot: using recombinant Tau isoforms, this antibody reacts with Tau isoforms containing four microtubule-binding motifs. On cortical brain tissue homogenates of Alzheimer disease patients, it specifically recognizes three hyperphosphorylated Tau components with an apparent molecular weight of 64, 69 and 74 kDa [3,4,5].
Cross-reaction: if used in excess, a cross reaction against other repeats is observed. It is important to use the antibody at the highest possible dilution, and to control the specificity of the labeling, using simultaneously the shortest recombinant tau (which should not be labeled) and the longest recombinant tau (which should be labeled).
2. Immunohistochemistry: this antibody is not specially good at the immunohitochemical level.
It could be improved by absorbing the serum with the shortest recombinant tau, or with homogenates of cells transfected with cDNA of tau without exon 10, or with a synthetic peptide corresponding the the third repeat (R3).
CONTROLS: the specificity should be controlled on brain tissue sections from Pick patients with Pick bodies and tangles (see Acta Neuropathol (Berl) 1994;87(2):115-24 Quantitative neuropathologic analysis of Pick's disease cases: cortical distribution of Pick bodies and coexistence with Alzheimer's disease.Hof PR, Bouras C, Perl DP, Morrison JH).
The good experimental conditions are those where tangles are labeled, but not Pick bodies.
Instructions for use:
1. For Western-blot, a final dilution of 1/1000 to 1/5000 (in Tris/NaCl/Tween buffer + 5% non-fat dry milk) for the detection of tau in tissue homogenates, and a staining with the ECL technique.
2. For immunohistochemistry, a final dilution of 1/500 (in Coons buffer) is suggested for immunostaining on frozen sections of cortical brain tissue.
Note: The recommended concentrations are approximative values. For each application, a dose response assay should be performed to determine the optimal concentration for use.FOR RESEARCH USE ONLY
References:
[1] Goedert M. and Jakes R., EMBO J. (1990); 9, 4225-4230.
[2] Goode B.L. and Feinstein S.C. J. Cell Biol. (1994): 124(5), 769-782.
[3] Delacourte et al., J. Neupathol. Exp. Neurol. (1996); 55, 159-168.
[4] Ann Neurol 1998 Feb;43(2):193-204
Vulnerable neuronal subsets in Alzheimer's and Pick's disease are distinguished by their tau isoform distribution and phosphorylation.
Delacourte A, Sergeant N, Wattez A, Gauvreau D, Robitaille Y
Unite INSERM 422, Lille, France.
Aggregated tau proteins constitute the basic matrix of neuronal inclusions specific to numerous neurodegenerative disorders. Monodimensional and two-dimensional Western blot analyses performed on cortical brain homogenates allowed discrimination between disease-specific tau protein profiles. These observations raised the issue of the physiopathological significance of such specificities. Alzheimer's disease (AD) pathological tau proteins (PTPs) (tau 74, 69, 64, 55) were compared with those of Pick's disease (PiD) (tau 64, 55) using a panel of antibodies against peptidic sequences of tau isoforms corresponding to exons 2, 3, and 10. AD and PiD could then be critically differentiated by the absence of translated tau isoforms with exon 10 in PiD PTPs, along with the absence of the phosphorylation site on Ser262. Immunohistochemical studies corroborate these findings. Indeed, Pick bodies were strongly immunostained by an anti-"exon 2" antibody but failed to reveal any anti-exon 10 reactive epitope. Tangles in AD contained exon 2, 3, and 10 epitopes. Altogether, our results demonstrated that Pick bodies develop within specific neuronal subsets that express specific patterns of 7 isoforms lacking exon 10 peptidic sequence. We conclude that neurodegenerative disorders imply attrition of selectively vulnerable neuronal subsets, a process revealed, and may be sustained by specific tau isoform patterns.
PMID: 9485060, UI: 98143594
[5] J Neurochem 1999 Mar;72(3):1243-9
Neurofibrillary degeneration in progressive supranuclear palsy and corticobasal degeneration: tau pathologies with exclusively "exon 10" isoforms.
Sergeant N, Wattez A, Delacourte A
INSERM Unite 422, Lille, France.
Pathological tau proteins that constitute the basic matrix of neuronal inclusions observed in numerous neurodegenerative disorders are disease specific. This is mainly the consequence of the aggregation of specific sets of tau isoforms according to the diseases, i.e., six isoforms in Alzheimer's disease (AD) and exclusively the three tau isoforms lacking the corresponding sequence of exon 10 (E10-) in Pick's disease (PiD). By using antibodies specific to the different tau isoforms and one- and two-dimensional gel electrophoresis followed by western blots, we demonstrate herein a third group of neurodegenerative disorders characterized by intraneuronal inclusions exclusively constituted of tau isoforms containing the sequence corresponding to exon 10, progressive supranuclear palsy (PSP) and corticobasal degeneration (CBD). Together, tau isoforms with exon 10 clearly differentiate three groups of neurodegenerative diseases: AD, PiD, and PSP/CBD. For each group, the neuropathological and clinical phenotypes are most likely related to specific sets of tau isoforms expressed by the vulnerable neuronal populations. The recently described mutations of the tau gene responsible for familial frontotemporal dementias also support this hypothesis.
PMID: 10037497, UI: 99155101
