Third Generation DNA Sequencing with a Nanopore - Nanopores: Sensing and Fundamental Biological Interactions - page 296

Fig. 12.3 Fluorescent images taken from a microfluidic device consisting of three channels that
merge into a single channel at a 15 angle. Fluorescent molecules indicate the flow lines. The
microfluidic is highlighted by white dashed lines. The flow is right to left in the channel. Four
different flow conditions are used to illustrate hydrodynamic focusing of fluorescent molecules:
(a) 0.03 m L/min; (b)10 m L/min; (c)5 m L/min; and (d) 2.5 m L/min

(from right to left) in a microfluidic device. By altering the ratio of the sheath (top
and bottom channels) to the input (center channel) flow rate, the flow in the center
channel can be focused into a thin jet - as evident from the fluorescent peak in the
center. Using this approach, we expect to convey single DNAs within the pore
capture radius without compromising the response time.
Alternatively, we designed a microfluidic chip with integrated micromechanical
valves enabling us to concentrate in the volume over the pore. This approach was
motivated by microfluidic applications such as genetic analysis of single cell
[ 53 , 54 ]. Figure 12.4a shows the plan of the chip, which we implemented with
soft lithography in polydimethylsiloxane (PDMS) using 12 m line-rules. This

design utilizes two-layer PDMS push-down microfluidic valves to control fluid
flow in a 10-input microchannel array that is bonded to a silicon nitride membrane
(either at high temperature

m

76 C for 3 h or using plasma oxidation). An elasto-
meric membrane is formed where the flow channel is positioned orthogonal to the
control channel directly above it. Magnified top-down views of two pressurized
valves are shown in Fig. 12.4c, d . According to this design, a single nanopore in a
membrane

>

m) 2 is located in the active volume at the intersection between
the horizontal and vertical microchannels, and independent fluid flows containing

< (10 m