Collagen IV Antibody Staining Protocol for Immunohistochemistry


Description: Collagen Type IV extracted and purified from mouse tumor tissues. Antibody shows less than 0.1% reactivity with human Collagen types IV, V and mouse Collagen Types I, II, III and mouse Fibronectin. It shows 4.0% reactivity with mouse Laminin by radioimmunoassay.


Primary Antibody

Name: Collagen IV Antibody

Clone: Rabbit anti-Mouse

Supplier: Chemicon

Catalog Number: AB756P

Dilution: 1:100 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 minutes/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 ºC - 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: Cytoplasmic
Images: Search image

Additional Information:
Tissue Type: Kidney
Fixation: Acetone or methyl-carnoy fixed paraffin-embedded sections or frozen sections.
Positive Control: Kidney
Negative Control: Omit primary antibody, isotype control or 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


No need to perform antigen retrieval on frozen sections