Journal: Biosensors & bioelectronics
Article Title: A Portable, Shock-Proof, Surface-Heated Droplet PCR System for Escherichia coli Detection
Figure Lengend Snippet: Schematic illustrations for the device layout and its operation. (A) All the major components of the device, less the circuit. A disposable cartridge, pre-loaded with solid coconut oil at room temperature and a droplet of PCR mixture (within the oil), is connected to the device. (B) The oil melts upon initial heating and the thermocouple loop picks up the droplet (PCR mixture + sample target). The sample solution is added to the PCR mixture droplet using a pipette. (C) In one complete thermal cycle, the droplet moves from the denaturation chamber (98°C), to the annealing chamber (50°C), and then to the extension chamber (80°C). The droplet returns back to the denaturation chamber to commence another cycle. The droplet is guided across the chambers by a thermocouple loop, and it contacts on the Teflon-coated heater surfaces. A PCB heater and a surface-mounted thermocouple control the oil temperature in each chamber. The droplet stays in each chamber until the thermocouple loop detects that it has reached the desired temperature (95°C, 56°C, and 72°C, respectively). (D) A pipette dislodges the droplet upon completion of PCR thermocycling. (E) The thermocouple loop and the metal guide are moved to the extension chamber to secure room for a smartphone microscope. 1 μL of 20× SYBR Green I dye solution is added to the droplet. (F) A smartphone-based fluorescence microscope measures fluorescence. Circuit layout as seen on the breadboards. (A) There are 3 MAX31855, one for each surface-mounted thermocouple. Three JZC-11F relays, one for each heater. The temperature and PID settings are displayed on a 20×4 serial LCD (not shown). (B) The motor controller circuit, showing the AD595 used for the thermocouple loop, which measures internal droplet temperature. Also shown is the EasyDriver connected to the Arduino microcontroller and Haydon-Kerk linear stepper motor. The output of the thermocouple is displayed on another 20×4 serial LCD (not shown). There are 3 buttons, one for starting thermocycling and two for manually positioning the thermocouple loop and droplet. Images created using Fritzing software (Friends of Fritzing e.V., Berlin, Germany). T/C =temperature control.
Article Snippet: Once the thermocycling is finished, 1 μL of 20× SYBR Green I (SG) dye, prepared from 10,000× SG (Molecular Probes – Life Technologies, Eugene, OR, USA), was added to the reaction droplet using a pipette, resulting in 2× SG in the final solution ( ).
Techniques: Polymerase Chain Reaction, Transferring, Microscopy, SYBR Green Assay, Fluorescence, Software