We report a practical world-to-chip microfluidic interfacing method with built-in valves suitable for microscale multichamber chip-based assays. One of the primary challenges associated with the successful commercialization of fully integrated microfluidic systems has been the lack of reliable world-to-chip microfluidic interconnections. After sample loading and sealing, leakage tests were conducted at 100 degrees C for 30 min and no detectable leakage flows were found during the test for 100 microchambers. To demonstrate the utility of our world-to-chip microfluidic interface, we designed a microscale PCR chip with four chambers and performed PCR assays. The PCR results yielded a 100% success rate with no contamination or leakage failures. In conclusion, we have introduced a simple and inexpensive microfluidic interfacing system for both sample loading and sealing with no dead volume, no leakage flow and biochemical compatibility.