Goat Anti-Rabbit IgG H&L (Biotin) (ab207995)

IHC image of Histone H4 staining in a section of formalin-fixed paraffin-embedded normal human colon tissue*. Ab207995 Goat Anti-Rabbit IgG H & L (Biotin) was used as the secondary antibody.

Staining was performed on a Leica Bond^TM. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins, before blocking of endogenous biotin using ab64212. The section was then incubated with ab177840, 1/100 dilution, for 15 mins at room temperature, followed by ab207995, 1/2000 dilution, for 15 mins at room temperature. Detection was via an HRP conjugated ABC system and DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

The inset negative control image is taken from an identical assay without primary antibody.

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.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

ab207995 was tested by direct ELISA, where wells were coated with serially diluted rabbit IgG (1000 – 16 ng/ml) for 2 hours, followed by a 2 hour blocking step (5% BSA). ab207995 (1:20,000 dilution; 2 hours) was added and detected by streptavidin-HRP (ab7403; 1:10,000 dilution; 1 hour). Signal was developed by TMB substrate. Data from duplicates; +/- SD.

IHC image of beta Tubulin staining in a section of formalin-fixed paraffin-embedded normal human colon tissue*. Ab207995 Goat Anti-Rabbit IgG H & L (Biotin) was used as the secondary antibody.

Staining was performed on a Leica Bond^TM. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins, before blocking of endogenous biotin using ab64212. The section was then incubated with ab6046, 1/100 dilution, for 15 mins at room temperature, followed by ab207995, 1/1000 dilution, for 15 mins at room temperature. Detection was via an HRP conjugated ABC system and DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

The inset negative control image is taken from an identical assay without primary antibody.

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.

*Tissue obtained from the Human Research Tissue Bank, supported by the NIHR Cambridge Biomedical Research Centre

Cross-reactivity of the polyclonal secondary antibody ab182016 was tested using a sandwich ELISA approach. The wells were coated with the indicated IgG standards at 1 µg/ml (50 µl/well) and incubated overnight at 4°C, followed by a 5% BSA blocking step for 2h at RT. ab182016 was then added starting at 1 µg/ml and gradually diluted 1/4 (50 µl/well), followed by incubation for 2h. For the detection Donkey anti-Goat IgG H&L (HRP) (ab6885) was used at 1/10,000 dilution (50 µl/well), followed by incubation for 1h at RT.

For the batch tested, ab182016 showed a cross-reactivity of 5-7% towards Human IgG and below 2% towards Mouse IgG, Rat IgG and Chicken IgY.

This data was developed using the unconjugated antibody (ab182016).

Cross-reactivity of Goat anti-Rabbit IgG H&L (ab182016) and Goat anti-Rabbit IgG H&L obtained from two different vendors was tested using a sandwich ELISA approach. The wells were coated with the indicated IgG standards (Rabbit, Human, Mouse and Rat) at 1 µg/ml (50 µl/well) and incubated overnight at 4°C, followed by a 5% BSA blocking step for 2h at RT. Secondary antibodies were then added starting at 1 µg/ml and gradually diluted 1/4 (50 µl/well), followed by incubation for 2h. For the detection Donkey anti-Goat IgG H&L (HRP) (ab6885) was used at 1/10,000 dilution (50 µl/well), followed by incubation for 1h at RT. This data is from a representative dilution.

This data was developed using the unconjugated antibody (ab182016).