Figure 1.
Microfluidic Chip housing three amplification chambers.
(A) An exploded view of the chip used for genotyping of malaria-transmitting mosquitoes. The chip consists of three layers: a top PMMA film; a PMMA chip body, and a PCR Sealers™ tape bottom. The various features of the chip body were milled with a CNC machine. (B) Cross-sectional view of the assembled, inverted chip loaded with a FTA disc. In the test chambers, the disks carry pieces of a mosquito leg. (C) A photograph of the chip.
Figure 2.
The sample preparation process.
(A) A mosquito leg was placed on the FTA® card. (B) The leg was crushed with a blunt object. Witness the resulting gray spot on the FTA® card. (C) A 1.5-mm diameter FTA disc was punched out from the spotted area with a Harris punch cutter.
Figure 3.
A schematic illustration of the experimental set-up for LAMP amplification and end-point detection.
The chip was heated by a thin-film heater to maintain the reactors at 63±0.5°C. At the conclusion of the LAMP amplification process, the chip was illuminated from the side with a blue LED and the photo of the fluorescent image was taken with a cell phone camera (iPhone 4).
Figure 4.
Real-time fluorescence amplification curves.
Real-time monitoring of LAMP amplification products in reactors G, A, and C as functions of time when the sample consists of An. gambiae (A) and An. arabiensis (B).
Figure 5.
End-point detection of An. gambiae DNA recorded with iPhone 4.
(A) An enlarged image of the illuminated chip. Reactor G emits visible light while reactors A and C stay dark. (B) Photograph of iPhone 4 with mosquito-on-a-chip fluorescence image. A minute, reusable, light-emitting diode (inset) excites the fluorescence emission.
Figure 6.
Electropherograms of LAMP products in a 2% agarose gel.
Lane M is DNA marker VIII. (A) An. gambiae identification. Lanes 1, 2, and 3 correspond, respectively, to LAMP products from reactors G, A, and C when the sample consists of An. Gambiae. (B) An. arabiensis identification. Lanes 1, 2, and 3 correspond, respectively, to LAMP products from reactors A, G, and C when the sample consists of An. arabiensis.