DNA Melting Analysis with Optofluidic Lasers Based on Fabry-Pérot Microcavity

Abstract

We conduct DNA high-resolution melting (HRM) analysis using optofluidic lasers based on a Fabry-Pérot microcavity. Compared to the fluorescence-based HRM, the laser-based HRM has advantages of higher emission intensity for better signal-to-noise ratio and sharper transition for better temperature resolution. In addition, the melting temperature can be lowered by optimizing the laser conditions such as external pump and cavity Q-factor. In this work, we first theoretically analyze the laser-based HRM. Then experiments are performed on three long DNA sequences as model systems, one being 99 bases and the other two being 130 bases long but with different GC contents. We show that the laser-based HRM is able to distinguish the target and the single-base mismatched DNA as long as 130 bases and with nearly 50% GC content. The dependence of laser threshold on the temperature for each DNA sample is first experimentally investigated and by optimizing the external pump, the melting temperature is reduced by more than 10 °C, compared to the fluorescence-based HRM for long DNA sequences up to 130 bases. Finally, we demonstrate an alternative method of using the laser-based HRM for rapid DNA screening that does not exist for the fluorescence-based HRM, in which laser excitation is scanned at a fixed temperature to distinguish the target and the base-mismatched DNA sequences. It is shown that the 130-bases-long DNA with nearly 50% GC content can have as much as 20% difference in the laser threshold and 40% difference in the laser output slope between the target and the single-base mismatched sequences, despite only 0.5 °C difference in their melting temperature, indicating that the laser-excitation-scanning method can also be suitable for long DNA sequences with higher GC content.

Fiber Optofluidic Microlasers: Structures, Characteristics, and Applications

“…The limit of detection is two orders of magnitude lower than that of fluorescence. [23] The detection of ions, [199,200] explosive, [201] uric acid, [202] and the DNA high-resolution melting analysis [203] have been demonstrated by optofluidic laser with a sensitivity much higher than traditional methods. Besides molecules, cellular level biodetection has been demonstrated by the cell-in-cavity or microcavity-in-cell strategies.…”

Biological Lasers for Biomedical Applications

“…Similar work was later carried out by Gong et al and Yang et al, showing the applications of the biolaser in sensitive sulfide ion detection (Figure b) and turbidimetric inhibition immunoassay, both of which had a larger dynamic range than the traditional methods. Besides immunoassay, Hou et al developed DNA high‐resolution melting (HRM) analysis by using the FP cavity based biolaser . Compared to the fluorescence‐based HRM, the laser‐based HRM has advantages of higher emission intensity for a better signal‐to‐noise ratio and a sharper transition for better temperature resolution.…”

Recent Advances of Organic Solar Cells with Optical Microcavities

“…MCs generally require at least one dimension of size in the order of magnitude of light wavelength. MC structure is also used in photodetectors, organic light emitting diodes (OLED), and lasers to improve their performance. Typical MC structures include Fabry–Perot (F–P) resonant cavity, whispering‐gallery mode (WGM), and so on.…”