A novel panel of α-synuclein antibodies reveal distinctive staining profiles in synucleinopathies.

Synucleinopathies are a spectrum of neurodegenerative diseases characterized by the intracellular deposition of the protein α-synuclein leading to multiple outcomes, including dementia and Parkinsonism. Recent findings support the notion that across the spectrum of synucleinopathies there exist diverse but specific biochemical modifications and/or structural conformations of α-synuclein, which would give rise to protein strain specific prion-like intercellular transmission, a proposed model that could explain synucleinopathies disease progression. Herein, we characterized a panel of antibodies with epitopes within both the C- and N- termini of α-synuclein. A comprehensive analysis of human pathological tissue and mouse models of synucleinopathy with these antibodies support the notion that α-synuclein exists in distinct modified forms and/or structural variants. Furthermore, these well-characterized and specific tools allow the investigation of biochemical changes associated with α-synuclein inclusion formation. We have identified several antibodies of interest with diverse staining and epitope properties that will prove useful in future investigations of strain specific disease progression and the development of targeted immunotherapeutic approaches to synucleinopathies

Summary of specificity and epitopes recognized by the novel αS antibodies.

<p>Summary of specificity and epitopes recognized by the novel αS antibodies.</p

Alignment of the amino- and carboxy-terminal amino acid sequences of human and mouse αS and compared to human βS and γS.

<p><b>(A)</b> Amino acid sequences of residues 2–21 in human αS, βS and γS and mouse αS. <b>(B)</b> Carboxy-terminal region sequence including amino acid residues 89–140 of human and mouse αS relative to the sequence of human βS and γS. Residues highlighted in orange indicate differences between human and mouse αS, while residues highlighted in yellow depict differences between human synuclein proteins.</p

Summary of the relative reactivity of αS inclusion pathology in human synucleinopathies and αS M83 mouse models with the new array of αS antibodies.

<p>Summary of the relative reactivity of αS inclusion pathology in human synucleinopathies and αS M83 mouse models with the new array of αS antibodies.</p

Representative immunohistochemistry of the cingulate gyrus of a DLB patient, and the cerebellum of a MSA patient stained with the indicated antibodies.

<p>Tissues sections from the cingulate cortex of a DLB patient (A) and the cerebellum of an MSA patient (B) were immunostained with each indicated antibody and developed with DAB as described in “Material and Methods and counterstained with hematoxylin. Arrows indicate cortical LBs in A and GCIs in B. Bar = 50 μm.</p

Alignment of the amino- and carboxy-terminal amino acid sequences of human and mouse αS and compared to human βS and γS.

<p><b>(A)</b> Amino acid sequences of residues 2–21 in human αS, βS and γS and mouse αS. <b>(B)</b> Carboxy-terminal region sequence including amino acid residues 89–140 of human and mouse αS relative to the sequence of human βS and γS. Residues highlighted in orange indicate differences between human and mouse αS, while residues highlighted in yellow depict differences between human synuclein proteins.</p

Representative immunohistochemistry depicting Lewy bodies stained with an array of αS antibodies within neuromelanin laden midbrain neurons of PD patients.

<p>Tissues sections were immunostained with each indicated antibody and developed with DAB as described in “Material and Methods” and counterstained with hematoxylin. Arrows indicate LBs. Bar = 50 μm.</p

Summary of the relative reactivity of αS inclusion pathology in human synucleinopathies and αS M83 mouse models with the new array of αS antibodies.

<p>Summary of the relative reactivity of αS inclusion pathology in human synucleinopathies and αS M83 mouse models with the new array of αS antibodies.</p

Summary of specificity and epitopes recognized by the novel αS antibodies.

<p>Summary of specificity and epitopes recognized by the novel αS antibodies.</p

Summary of M83 transgenic mice used for immunohistochemistry studies.

<p>Summary of M83 transgenic mice used for immunohistochemistry studies.</p