Immunohistochemical Detection of Retinoblastoma Protein Phosphorylation in Human Tumor Samples

The retinoblastoma protein (pRb) is an important tumor suppressor and cell cycle repressor. pRb is a phosphoprotein whose function is regulated primarily at the level of phosphorylation, and therefore, detecting pRb's phosphorylation status in human tissue samples can be clinically informative. Unfortunately, detection of phosphorylated pRb residues can be technically challenging, as these residues can often be weak antigens. In this chapter, we describe an enhanced sensitivity immunohistochemistry protocol for the staining of phosphorylated serine 249 in pRb, in human lung tumor samples.