An Immunofluorescence Assay to Detect Alpha-Synuclein Aggregates in a Skin Biopsy Section

Published: April 30, 2024

Abstract

Source: Vacchi, E., et al. Targeting Alpha Synuclein Aggregates in Cutaneous Peripheral Nerve Fibers by Free-floating Immunofluorescence Assay. J. Vis. Exp. (2019).

This video demonstrates an immunofluorescence assay to detect neurodegenerative disease-specific alpha-synuclein aggregates on skin biopsy sections. Free-floating immunostaining is employed to fluorescently label the carboxyl-terminal hydrolase and the alpha-synuclein aggregates in the dermal neurons. Under a confocal microscope, cells with fluorescence from both antibodies indicate the presence of alpha-synuclein aggregates in the dermal neurons.

Protocol

All procedures involving sample collection have been performed in accordance with the institute's IRB guidelines, 1. Skin Biopsy Collection Let a qualified physician perform the skin biopsy in an appropriate clinical setting. Choose the area to perform the skin biopsy and clean it with an alcohol swab. Prepare the anesthetic solution with 1 cc of lidocaine 2%. With the needle parallel to the area, inject local anesthesia subcutaneo…

Divulgazioni

The authors have nothing to disclose.

Materials

5G4 (anti human αSyneclein 5G4) Analytik Jena Roboscreen 847-0102004001 Mouse monoclonal
AlexaFluor 488 Goat anti Rabbit IgG Invitrogen 1971418 2mg/ml
AlexaFluor 594 Goat anti Mouse IgG Invitrogen 1922849 2mg/ml
Disodium hydrogen phosphate solution Merk Millipore 106586
Ethylene Glycol Sigma-Aldrich 324558
Glycerol Sigma-Aldrich G7757
L-Lysine monohydrochloride Sigma-Aldrich L5626
Paraformaldehyde Aldrich Chemistry 441244
PGP9.5 Abcam ab15503 Rabbit polyclonal
Sodium Chloride Sigma S3014
Sodium Dihydrogen Phosphate Monohydrate Merck Millipore 106346
Sodium (meta)periodate Sigma-Aldrich S1878
Trizma Base Sigma T1503
Tryton X-100 Sigma-Aldrich X100
Vectashield Vector Laboratories H-1000 Mounting medium

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Citazione di questo articolo
An Immunofluorescence Assay to Detect Alpha-Synuclein Aggregates in a Skin Biopsy Section. J. Vis. Exp. (Pending Publication), e22111, doi: (2024).

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