T echniques and Methods Affinity Detection and Quantification of Receptors of TRAIL Based on Their Thermostability

Objective To develop a way for tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptors quantification in cancer via its’ thermostability. Methods Endougenous alkaline phosphatase (AP) activity and denaturation temperature of pancreatic cancer cell lines AsPC-1 and Capan-2 were detected. Boiling treated recombinant protein death receptor 5 (DR5) ,named DR5-AP, as well as pancreatic cancer cells lines AsPC-1 and Capan-2 were incubated with AP-tagged TRAIL (AP-TRAIL), and then reacted with Reagent A and Reagent S, the substrate of AP, to quantitive and in site detection of the receptor. Results The endougenous AP activity of pancreatic cancer cells lines AsPC-1 and Capan-2 could not be totally inactivated by incubated at 65 ℃, thus inhibited the detection of TRAIL receptor, but the activity was dramatically decreased after treated with boiling water, whereas the DR5-AP was thermal stable. The surface receptor of AsPC-1 and Capan-2 could be recognized and bound by AP-TRAIL after treated at 100 ℃, the readings were 2.210±0.393 and 2.027±0.019. Conclusion The TRAIL receptors are thermostable and this may provide a better diagnosis and prognosis of cancer as well as personalize cancer therapy.