Immunogold labelling of paired helical filaments and amyloid fibrils by specific monoclonal and polyclonal antibodies

Reig, Sylviane Hennebico; Buée‐Scherrer, Valérie; Mourton-Gilles, Chantal; Défossez, A.; Delacourte, André; Beauvillain, Jean-Claudé; Mazzuca, M

doi:10.1007/bf00294803

Cited by 11 publications

(5 citation statements)

References 38 publications

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“…AD2 is a protein A‐purified monoclonal antibody raised against a crude PHF preparation obtained from AD brain [23]. It detects NFTs by light microscopy [5], PHFs by electron microscopy [24]and PHF‐Tau on immunoblots [23]. It specifically recognizes phosphorylated serines 396 and 404 (numbering according to the longest human brain Tau isoform) in the carboxy‐terminal part of Tau [23].…”

Section: Methodsmentioning

confidence: 99%

Different distribution of phosphorylated tau protein isoforms in Alzheimer's and Pick's diseases

et al. 1997

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Section: Methodsmentioning

confidence: 99%

Different distribution of phosphorylated tau protein isoforms in Alzheimer's and Pick's diseases

et al. 1997

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“…Since 1971, gold NPs conjugated with a specific antibody have been successfully used to label amyloids (32). This technique, called immunogold labeling, allows for the localization of amyloids in tissue sections and the characterization of the sequence and structural properties of fibrils in solution (33,34). However, with this technique, NPs do not anchor directly on the fibrils, but are located as far as 30 nm away due to the presence of the bulky antibody and linker.…”

Section: Mus:ot Nps Successfully Label Different Polymorphs Of Amyloidmentioning

confidence: 99%

Unraveling the complexity of amyloid polymorphism using gold nanoparticles and cryo-EM

Cendrowska

Silva

Ait-Bouziad

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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Increasing evidence suggests that amyloid polymorphism gives rise to different strains of amyloids with distinct toxicities and pathology-spreading properties. Validating this hypothesis is challenging due to a lack of tools and methods that allow for the direct characterization of amyloid polymorphism in hydrated and complex biological samples. Here, we report on the development of 11-mercapto-1-undecanesulfonate-coated gold nanoparticles (NPs) that efficiently label the edges of synthetic, recombinant, and native amyloid fibrils derived from different amyloidogenic proteins. We demonstrate that these NPs represent powerful tools for assessing amyloid morphological polymorphism, using cryogenic transmission electron microscopy (cryo-EM). The NPs allowed for the visualization of morphological features that are not directly observed using standard imaging techniques, including transmission electron microscopy with use of the negative stain or cryo-EM imaging. The use of these NPs to label native paired helical filaments (PHFs) from the postmortem brain of a patient with Alzheimer’s disease, as well as amyloid fibrils extracted from the heart tissue of a patient suffering from systemic amyloid light-chain amyloidosis, revealed a high degree of homogeneity across the fibrils derived from human tissue in comparison with fibrils aggregated in vitro. These findings are consistent with, and strongly support, the emerging view that the physiologic milieu is a key determinant of amyloid fibril strains. Together, these advances should not only facilitate the profiling and characterization of amyloids for structural studies by cryo-EM, but also pave the way to elucidate the structural basis of amyloid strains and toxicity, and possibly the correlation between the pathological and clinical heterogeneity of amyloid diseases.

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“…Since 1971, gold NPs conjugated with a specific antibody have been successfully used to label amyloids (33). This technique, called immunogold labeling, allows for the localization of amyloids in tissue sections and characterization of the sequence and structural properties of fibrils in solution (34,35). However, with this technique, NPs do not anchor directly on the fibrils but are located as far as 30 nm away due to the presence of the bulky antibody and linker.…”

Section: Mus:ot Nps Successfully Label Different Polymorphs Of Amyloimentioning

confidence: 99%

Unraveling the Complexity of Amyloid Polymorphism Using Gold Nanoparticles and Cryo-EM

Cendrowska

Silva

Ait-Bouziad

et al. 2019

Preprint

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The ability of proteins to self--assemble into different types of fibrils with distinct morphologies has been linked to the pathological and clinical heterogeneity of amyloid diseases such as Alzheimer's and Parkinson's disease. Herein, we describe novel nanoparticles (NPs) that efficiently label amyloid fibrils produced in vitro or isolated from postmortem tissues, under hydrating conditions and in such a way to unmask their polymorphism and morphological features. Using these NPs, we show that pathological aggregates exhibit exceptional morphological homogeneity compared to amyloid fibrils produced in vitro, consistent with the emerging view that the physiologic milieu is a key determinant of amyloid fibril strains. These advances pave the way for elucidating the structural basis of amyloid strains and toxicity. AbstractThe misfolding and self--assembly of proteins into β--sheet--rich amyloid fibrils of various structures and morphologies is a hallmark of several neurodegenerative and systemic diseases. Increasing evidence suggests that amyloid polymorphism gives rise to different strains of amyloids with distinct toxicity and pathology--spreading properties. Validating this hypothesis is challenging due to a lack of tools and methods that allow for the direct characterization of amyloid polymorphism in hydrated and complex biological samples. Here, we report on the use of 11--mercapto--1--undecanesulfonate--coated gold nanoparticles (NPs) to label the edges of synthetic, recombinant and native amyloid fibrils to assess amyloid morphological polymorphism using cryogenic transmission electron microscopy (cryo--EM).The fibrils studied were derived from amyloid proteins involved in disorders of the central nervous system (amyloid--β, tau, α--synuclein) and in systemic amyloidosis (a fragment of an immunoglobulin λ light chain). The labeling efficiency enabled imaging and characterization of amyloid fibrils of different morphologies under hydrated conditions using cryo--EM. These NPs allowed for the visualization of morphological features that are not directly observed using standard imaging techniques, including TEM with use of the negative stain or cryo--EM imaging. We also demonstrate the use of these NPs to label native paired helical filaments (PHFs) from the postmortem brain of an Alzheimer's disease patient, as well as amyloid fibrils extracted from the heart tissue of a patient suffering from systemic amyloid light--chain (AL) amyloidosis. Analysis of the cryo--EM images of amyloids decorated with NPs shows

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Immunogold labelling of paired helical filaments and amyloid fibrils by specific monoclonal and polyclonal antibodies

Cited by 11 publications

References 38 publications

Different distribution of phosphorylated tau protein isoforms in Alzheimer's and Pick's diseases

Different distribution of phosphorylated tau protein isoforms in Alzheimer's and Pick's diseases

Unraveling the complexity of amyloid polymorphism using gold nanoparticles and cryo-EM

Unraveling the Complexity of Amyloid Polymorphism Using Gold Nanoparticles and Cryo-EM

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