A highly sensitive bioassay for PTH using ROS 17/2.8 subclonal cells

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Abstract
A highly sensitive bioassay for PTH was developed by using rat osteosarcoma cells (ROS 17/2.8). By limiting dilution, ROS cells were subcloned and the subclonal cell line (ROS 17/2.8–5) most responsive to PTH was selected. When subconfluent ROS 17/2.8–5 cells were treated with hydrocortisone for 3 days and then incubated with PTH, the cAMP response was significant at 10–40 ng/l hPTH (1–34) (4 ∼ 16 × 10−12 mol/l). Osteoclast activating factors such as human interleukin I alpha and beta, and tumour necrosis factor alpha did not stimulate cAMP production, whereas a conditioned medium of oesophageal carcinoma cells established from a patient with humoral hypercalcaemia stimulated cAMP production. By selecting PTH-responsive subclonal cells and treating them with hydrocortisone, the sensitivity for detecting PTH was improved approximately 15 times. This method will be useful in the characterization and purification of PTH-like factors produced by malignant tumours from hypercalcaemic patients.