Recombinant Biotin Anti-Human Kappa Chain antibody [EPR5539-71-8] (ab201490)

Ab201490 was tested using a sandwich ELISA approach. The wells were coated with ab195576 at 0.5µg/ml at 50µl/well overnight at 4°C, followed by a 5% BSA blocking step for 2h RT. Human IgG (ab98981) was then added starting at 40 µg/ml and plasma/serum at 1:50 and gradually diluted 1:4, 50µl/well for 2h. Ab20149 was then added at 1:10,000 dilution, 50µl/well for 2h. A HRP-streptavidin (ab7403) was used at 1:10,000 dilution for 1h.

IHC image of human kappa chain staining in a section of formalin-fixed paraffin-embedded normal human tonsil*, performed on a Leica Bond system using the standard protocol B. 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 ab201490, at 1/100 dilution, for 15 mins at room temperature and detected using an HRP conjugated ABC system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

Staining corresponds to B cells expressing IgG commonly found in lymphoid follicles of human tonsil.

The inset background control image is taken from an identical assay without added 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

Negative IHC image of human kappa chain staining in a section of formalin-fixed paraffin-embedded normal human cerebral cortex*, performed on a Leica Bond system using the standard protocol B. 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 ab201490, at 1/100 dilution, for 15 mins at room temperature and detected using an HRP conjugated ABC system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.

Staining corresponds to immunoglobulins found in serum. As expected, staining diminishes further into the brain tissue due to the action of the blood-brain barrier.

The inset background control image is taken from an identical assay without added 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