RhoH Colorimetric Cell-Based ELISA

From Assay Genie, this Colorimetric Cell-Based ELISA is a sensitive, lysate-free, high-throughput assay designed to detect the expression profile of the RhoH protein in cultured cells. The kit targets RhoH, a small GTPase of the Ras superfamily that acts as a negative regulator of hematopoietic cell proliferation and a critical mediator of T-cell antigen receptor signaling and mast cell activation, and is positioned as a relevant tool for studies of immune regulation and diseases such as autoimmune disorders and cancer.

The assay is reported reactive with Human, Mouse, and Rat samples and uses an indirect ELISA format in which RhoH-specific primary antibodies are detected by HRP-conjugated secondary antibodies, producing a colorimetric reaction read at 450 nm. The dynamic range is > 5000 Cells, the format is a 96-Well Microplate suitable for adherent cell culture, and storage is at 4 °C with 6 months stability.

A monoclonal anti-GAPDH antibody is included as an internal positive control for normalization of target absorbance, and a Crystal Violet whole-cell staining step is provided to normalize results to cell density. The standard procedure seeds 200 µL of 20,000 adherent cells per well, with a Poly-L-Lysine coating option for suspension cells, and proceeds through fixation, quenching, blocking, overnight primary antibody incubation, secondary antibody incubation, substrate addition, and stop solution before the absorbance is read at 450 nm.