Alpha-1-Antichymotrypsin Antibody Staining Protocol for Immunohistochemistry

Alpha 1 Antichymotrypsin Antibody Staining Protocol for Immunohistochemistry

Description: Alpha-1-Antichymotrypsin (ACT) is an early-stage acute phase protein and a member of the serine proteinase inhibitor or serpin superfamily. The precise role of ACT is uncertain but it is thought to act as an anti-inflammatory agent inhibiting chymotrypsin, cathepsin G, mast cell chymase, neutrophil chemotaxis and superoxide anion production. ACT is synthesized primarily by hepatocytes of the liver. Lower levels of synthesis have also been discovered via immunohistochemical analysis in mast cells, endothelial cells, breast and intestinal epithelial cells. ACT also exists in the brain. Research has shown that it is found in amyloid fibrils, endothelial cells and the cytoplasm of astroglial cells in certain brain abnormalities. Further research has also shown that a major proportion of prostate-specific antigen (PSA) in serum exists complexed to ACT.

Primary Antibody

Name: alpha-1-Antichymotrypsin Antibody

Clone: Rabbit Polyclonal

Supplier: Novocastra Labs

Catalog Number: NCL-A1ACp

Dilution: 1:100 - 1:200 using IHC-Tek^TM 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: Antigen retrieval is not needed

Buffer/pH value: N/A

Heat/Cool Temperature: N/A

Heat/Cool Time: N/A

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

Counterstain

Reagent: Mayer's Hematoxylin

Staining Time: 30 seconds

Results:

Staining Pattern: Cytoplasmic

Images: Search image

Additional Information:

Tissue Type: Tonsil, placenta

Fixation: Formalin-fixed paraffin sections, Not effective on frozen sections

Positive Control: Tonsil, placenta

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

References:

1. Niki T, Oka T, Shiga J, Machinami R (1995) Kupffer cells in multiple organ failure--their activation as revealed by immunohistochemistry for lysozyme, alpha 1-antichymotrypsin, and lectins. Gen Diagn Pathol. 141(1):21-7 PubMed Abstract

2. Uchida K, Kuroki K, Yoshino T, Yamaguchi R, Tateyama S. (1997)Immunohistochemical study of constituents other than beta-protein in canine senile plaques and cerebral amyloid angiopathy.
Acta Neuropathol (Berl). 93(3):277-84. PubMed Abstract