IHH and osteosarcoma: Since there are currently no specific antibodies for endogenous human Hedgehog pathway genes that work reliably in immunohistochemistry assays [27], the transcript levels of IHH, SMO, PTCH1, and GLI1 were determined using real-time RT-PCR in 43 human primary high-grade osteosarcoma samples and normalized to the levels detected in normal osteoblasts (Figure 1).