Iba1 Antibody Staining Protocol for Immunohistochemistry

Description: This antibody is expected to recognise the isoforms represented by NP_001614 and NP_116573 but not NP_004838 Please note: Although we have some very good Abreviews on mouse, some customers were receiving inconsistent results on mouse samples. We have therefore moved mouse to the predicted species and can longer guarantee it.

Primary Antibody

Name: Iba1 Antibody

Clone: Goat polyclonal

Supplier: Abcam

Catalog Number: ab5076

Dilution: 1:100 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: IHC-Tek^TM Epitope Retrieval Steamer Set (Cat# IW-1102)

Buffer/pH value: IHC-Tek^TM Epitope Retrieval Solution (Cat# IW-1100)

Heat/Cool Temperature: 95-100 ºC/room temperature

Heat/Cool Time: 20 minutes/20 minutes

This antibody did not work on citrate buffer based antigen retrieval solution.

Detection Methods

Standard Method: ABC Method or LSAB Method

Enhanced Method: Polymeric Methods

Chromogen Substrate

Reagent: DAB Chromogen Substrate

Incubation Time/Temperature: 1-3 minutes/room temperature

Counterstain

Reagent: Mayer's Hematoxylin

Staining Time: 30 seconds

Results:

Staining Pattern: Cytoplasm > cytoskeleton. Cell projection > ruffle membrane. Associated with the actin cytoskeleton at membrane ruffles and at sites of phagocytosis.

Images: Search image

Additional Information:

Species Reactivity: Rat, guinea pig, human, pig, mouse

Fixation: Formalin fixed paraffin sections

Positive Control: Brain

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: