Alpha Synuclein Antibody Staining Protocol for Immunohistochemistry


Description: Alpha-synuclein is a protein of 140 amino acids and a member of the synuclein family. It shares sixty one per cent of its sequence with beta-synuclein and is highly conserved

between vertebrate species. It does not possess a signal sequence suggesting that it is an

intracellular protein. All synucleins have the unusual organisation based around an eleven residue repeating motif and an alpha-helical secondary structure resembling those found in the lipid-binding domain of exchangeable apolipoproteins including Apo E. This homology suggests a direct interaction of alpha-synuclein with membranes consistent with its affinity for synaptosomes. The function of alpha-synuclein may be to carry a target protein to the inner membrane of nerve terminals or to the outer surface of synaptic vesicles.


Primary Antibody

Name: alpha-Synuclein Antibody

Clone: KM51, Mouse

Supplier: Novocastra Labs

Catalog Number: NCL-ASYN

Dilution: 1:20 - 1:40 using IHC-TekTM Antibody Diluent (Cat# IW-1000 or IW-1001) to reduce background and unspecific staining and serum blocking step is NOT needed.

Incubation Time/Temp: 60 min/room temperature

Antigen Retrieval
Device: IHC-TekTM Epitope Retrieval Steamer Set (Cat# IW-1102)
Buffer/pH value: IHC-TekTM Epitope Retrieval Solution (Cat# IW-1100)
Heat/Cool Temperature: 95-100 ºC/room temperature
Heat/Cool Time: 20 minutes/20 minutes

Detection Methods
Standard Method: ABC Method or LSAB Method
Enhanced Method: Polymeric Methods

Chromogen Substrate
Reagent: DAB
Incubation Time/Temperature: 1-3 minutes/room temperature

Reagent: Mayer's Hematoxylin
Staining Time: 30 seconds

Staining Pattern: Lewy body and neuropil staining
Images: Search image

Additional Information:
Tissue Type: Posterior cingulated gyrus (Lewy body dementia).
Fixation: Formalin-fixed paraffin sections
Positive Control: Posterior cingulated gyrus (Lewy body dementia).
Negative Control: Omit primary antibody, isotype control, absorption control
Blocking: 2-5% normal serum to reduce unspecific background staining; 0.5-3% H2O2 to block endogenous peroxidase activity; avidin/biotin to block endogenous biotin activity if necessary