Demonstration of the Activity of P-glycoprotein by a Semi-automated Fluorometric Method

作者：Spengler Gabriella; Viveiros Miguel; Martins Marta; Rodrigues Liliana; Martins Ana; Molnar Jozsef; Couto Isabel; Amaral Leonard^*

来源：Anticancer Research, 2009, 29(6): 2173-2177.

• ABC transporters
• P-glycoprotein
• semi-automated fluorometric method
• efflux
• ethidium bromide
• mouse lymphoma cells
• mdr1
• ABCB1

Background: We have developed a semi-automated fluorometric method that utilizes ethidium bromide (EB), a common substrate of bacterial efflux pumps. The method is sufficiently sensitive to characterize the efflux pump systems of bacteria. Because EB is also recognized and extruded by ATP-binding cassette (ABC) transporters and these have similarity to P-glycoprotein (P-gp), the method has been extended for the evaluation of agents that can inhibit the extrusion of EB on a real-time basis by mouse lymphoma cells containing the human ABCB1 (mdr1) gene. Materials and Methods: Monitoring of uptake and extrusion of EB was assessed using the Rotor-Gene(TM) 3000 (Corbett Research) under different conditions. Results: Whereas extrusion of EB took place readily, the addition of known inhibitors of efflux pumps (verapamil, reserpine) caused retention of EB. Conclusion: This method is inexpensive and allows the detection of neoplastic cells with increased efflux activity as well as the screening of large numbers of compounds for inhibition of the P-gp.

• 出版日期2009-6
• 单位河北医科大学