Original Paper

Acta Neuropathologica

, 116:277

First online:

Evaluation of α-synuclein immunohistochemical methods used by invited experts

  • Thomas G. BeachAffiliated withCivin Laboratory for Neuropathology, Sun Health Research Institute Email author 
  • , Charles L. WhiteAffiliated withNeuropathology Laboratory and Pathology Immunohistochemistry Laboratory, University of Texas Southwestern Medical School
  • , Ronald L. HamiltonAffiliated withDepartment of Pathology, University of Pittsburgh School of Medicine
  • , John E. DudaAffiliated withParkinson’s Disease Research, Education and Clinical Center, Philadelphia Veterans Affairs Medical Center
  • , Takeshi IwatsuboAffiliated withDepartment of Life-Pharmaceutics, University of Tokyo
  • , Dennis W. DicksonAffiliated withDepartment of Laboratory Medicine and Pathology, Mayo Clinic
  • , James B. LeverenzAffiliated withDepartment of Neurology, University of WashingtonDepartment of Psychiatry and Behavioral Sciences, University of WashingtonVeterans Affairs Puget Sound Health Care System
  • , Federico RoncaroliAffiliated withDepartment of Clinical Neuroscience, Imperial College
  • , Manuel ButtiniAffiliated withElan Pharmaceuticals
    • , Christa L. HladikAffiliated withNeuropathology Laboratory and Pathology Immunohistochemistry Laboratory, University of Texas Southwestern Medical School
    • , Lucia I. SueAffiliated withCivin Laboratory for Neuropathology, Sun Health Research Institute
    • , Joseph V. NoorigianAffiliated withParkinson’s Disease Research, Education and Clinical Center, Philadelphia Veterans Affairs Medical Center
    • , Charles H. AdlerAffiliated withDepartment of Neurology, Mayo Clinic

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Abstract

The use of α-synuclein immunohistochemistry has altered our concepts of the cellular pathology, anatomical distribution and prevalence of Lewy body disorders. However, the diversity of methodology between laboratories has led to some inconsistencies in the literature. Adoption of uniformly sensitive methods may resolve some of these differences. Eight different immunohistochemical methods for demonstrating α-synuclein pathology, developed in eight separate expert laboratories, were evaluated for their sensitivity for neuronal elements affected by human Lewy body disorders. Identical test sets of formalin-fixed, paraffin-embedded sections from subjects diagnosed neuropathologically with or without Lewy body disorders were stained with the eight methods and graded by three observers for specific and nonspecific staining. The methods did not differ significantly in terms of Lewy body counts, but varied considerably in their ability to reveal neuropil elements such as fibers and dots. One method was clearly superior for revealing these neuropil elements and the critical factor contributing to its high sensitivity was considered to be its use of proteinase K as an epitope retrieval method. Some methods, however, achieved relatively high sensitivities with optimized formic acid protocols combined with a hydrolytic step. One method was developed that allows high sensitivity with commercially available reagents.