In this paper, we present a novel multiple-stage amplifier chain based strong physical unclonable function (PUF) with low power and energy consumption. Based on the proposed two-dimensional subthreshold amplifier array, 12 different amplifiers can be selected through the analog multiplexer at each column. As a result, a 12-stage amplifier chain can be formed by applying different challenges to the aforesaid analog multiplexers with a linear feedback shift register (LFSR). Due to the inevitable process variation, the output voltage of the amplifier chain’s last stage varies depending on the various combinations of the selected amplifiers, whose number features an exponential relationship with the size of the adopted amplifier array. By using 65nm standard CMOS process, the proposed strong PUF implementation is validated with high reliability and randomness. According to our extensive simulation results, the averaged bit error rate (BER) per 10◦C and BER per 0.1V are calculated to be 3.15% and 3.85% for the operating temperature range of −20◦C∼120◦C and supply voltage range of 0.9V∼1.4V, respectively. Meanwhile, the proposed strong PUF’s high randomness is also verified by passing both the NIST and auto-correlation function (ACF) test suites. Moreover, featuring an excellent uniqueness of 49.54%, the overall power consumption is simulated to be 0.128µW at the throughput of 0.02Mb/s, which corresponds to an energy consumption as low as 6.4pJ/bit.