Loop-mediated isothermal amplification (LAMP) is a novel nucleic acid amplification method. Compared with the widely utilized polymerase chain reaction (PCR), LAMP has higher speed and efficiency as well as lower requirement for system temperature control because the whole amplification process is isothermal and no efforts are needed to switch between different temperatures. In this paper, we designed and fabricated different kinds of polycarbonate (PC) microfluid chips, explored appropriate reaction condition for LAMP in microenvironment (1 nL→10 μL), and developed a microfluidic isothermal amplification detection system. The DNA optimal amplification temperature is obtained; the starting time of exponential amplification of DNA is put forward farther. The optimal condition of DNA amplification in microenvironment, with a little reaction materials and early starting exponential amplification time of DNA are very important for clinic DNA detection and the application of Lab-on-a-Chip.