(A) RT-PCR detection of RET and GDNFR-α mRNA in primary BMSC from two donors (nos. 1 and 2), BMF, Saos-2, and MG-63 osteoblast cell lines and C433 stromal cell line. Aliquots of cDNA bulks were amplified with primer pairs specific for RET and GDNFR-α, run in agarose gels, blotted, and hybridized with oligoprobes specifically designated to recognize PCR-amplified products. An adult human brain substantia nigra cDNA and cDNA derived from the MN-60 cell line were used, respectively, as positive (+) and negative (−) controls for RET and GDNFR-α expression. cDNAs were always tested with β-actin–specific primers. (B) Fluorescence histograms showing expression of surface GPI-linked GDNF receptors (GDNFR-α) in BMSC, BMF, and the MG-63 cell line. Cells pretreated or not with PI-PLC (1 U/mL for 1 hour at 37°C) were incubated for 1 additional hour with GDNF (100 ng/mL) and then sequentially with chicken polyclonal antihuman GDNF (100 μg/mL) and rabbit fluorescein isothiocyanate-conjugated isotype-matched antichicken Igs. As controls, isotype-matched irrelevant chicken Igs were used.