Immunohistochemical analysis of adult mouse testis tissue, staining Oligodendrocyte Specific Protein (red) with ab7474.

Tissue was blocked with 5% BSA for 1 hour at 25°C. Samples were incubated with primary antibody (1/100 in 1% BSA) for 1 hour at 25°C. A TRITC-conjugated donkey anti-rabbit polyclonal IgG (1/200) was used as the secondary antibody.

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Immunofluorescent staining for OSP obtained with OSP antibody (ab7474) in rat brain cortex. The picture shows oligodendrocyte immunostaining in the cingular cortex. Picture taken with X20 objective, cells stained are ~30microns in diameter. Animals were intracardially perfused with 4% PFA. Tissue was post-fixed overnight in the same fixative, cryoprotected in 20% sucrose and frozen in OCT. Protocol : free floating IHC on 30um cryostat sections. Primary antibody ab7474 was used at 1/100 and incubated overnight at RT. Secondary antibody, Alexa fluor 488 at 1/1000, incubated for 2h at RT.

IHC image of ab7474 staining in human hippocampus formalin fixed paraffin embedded tissue section, performed on a Leica Bond^TM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab7474, 5µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

For other IHC staining systems (automated and non-automated) customers should optimize variable parameters such as antigen retrieval conditions, primary antibody concentration and antibody incubation times.