Miniaturized PCR chips for nucleic acid amplification and analysis: latest advances and future trends

Zhang, Chunsun; Xing, Da

Semi-invasive temperature measurement based on a fluorescence dye indicator. The principle of this temperature measuring technique has been described by Sakakibara and Adrian (95) . Although this technique has been well studied, it was not applied for the temperature measurement within the PCR chips until recently. Guttenberg et al. used a thermochrome dye chromazone-based temperature measuring technique to yield more temperature information inside the PCR sample droplet (77). Neuzil et al. investigated the temperature measurement of the on-chip PCR sample in SYBR Green-based system or 6-FAM-based system (80) . SYBR Green allows melting curve analysis (MCA) to be routinely used for PCR sample temperature monitoring, but it can be used with high precision only around the DNA melting point. By contrast, the fluorescence intensity of 6-FAM is proportional to its temperature up to 908C and thus it is more suitable to estimate the PCR sample temperature over a wide range. Most recently, rhodamine B, one of most widely used temperature sensitive dyes, has been utilized to evaluate the accuracy of the in-channel temperature measured by K-type thermocouple or by numerical simulation (39) . This dye-based measuring technique has advantages of simplicity, a wide range of temperature measurement, fast response speed, good spatial resolution and great sensitivity. However, its measurement accuracy is low.