RESUMO
We developed an integrated chip for real-time amplification and detection of nucleic acid using pH-sensing complementary metal-oxide semiconductor (CMOS) technology. Here we show an amplification-coupled detection method for directly measuring released hydrogen ions during nucleotide incorporation rather than relying on indirect measurements such as fluorescent dyes. This is a label-free, non-optical, real-time method for detecting and quantifying target sequences by monitoring pH signatures of native amplification chemistries. The chip has ion-sensitive field effect transistor (ISFET) sensors, temperature sensors, resistive heating, signal processing and control circuitry all integrated to create a full system-on-chip platform. We evaluated the platform using two amplification strategies: PCR and isothermal amplification. Using this platform, we genotyped and discriminated unique single-nucleotide polymorphism (SNP) variants of the cytochrome P450 family from crude human saliva. We anticipate this semiconductor technology will enable the creation of devices for cost-effective, portable and scalable real-time nucleic acid analysis.