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

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 approximately 16 X 10(-12) mol/l). Osteoclast activating factors such as human interleukin 1 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.