Chapter 7 – Complementary Experimental Tools  295

away from the mold, trimmed, and, if appropriate, bonded to a glass coverslip by drying the

PDMS and subjecting both the PDMS and coverslip to plasma cleaning and then simply

pressing the two surfaces together.

In this way, several complex, bespoke flow-​cell designs can be generated (Figure 7.5).

These enable biological samples to be immobilized in the sample chamber and observed con­

tinuously over long time scales (from minutes to several days if required) using light micros­

copy techniques. An important application uses multichannel inputs, which enables the fluid

environment of the same biological sample (e.g., a collection of immobilized living cells on

the microscope coverslip surface) to be exchanged rapidly typically in less than ~1 s. This

has significant advantages in enabling observation of the effects of changing the extracellular

environment on the exact same cells and in doing so circumvents many issues of cell-​to-​

cell variability in a cell population that often makes definitive inference more challenging

otherwise.

Microfluidics is also used in several high-​throughput detection techniques, including

FACS (discussed in Chapter 3). A more recent application has been adapted to traditional

PCR methods (see the previous section of this chapter). Several commercial microfluidics

PCR devices can now utilize microliter volumes in parallel incubation chambers. This can

result in significant improvements in throughput. This general microfluidics-​driven approach

of reducing sample incubation volumes and parallelizing/​multiplexing these volumes shows

promise in the development of next-​generation sequencing techniques, for example, in

developing methods to rapidly sequence the DNA from individual patients in clinics and all

parts of important progress toward greater personalized medicine (discussed in Chapter 9).

Using microfluidics, it is now possible to isolate individual cells from a population, using

similar fluorescence labeling approaches as discussed previously for FACS (see Chapter 3)

and then sequence the DNA from that one single cell. This emerging technique of single-​cell

FIGURE 7.5  Microfluidics. PDMS can be cast into a variety of microfluidics flow-​cell designs

using a solid substrate silicon-​based mask manufactured using microfabrication techniques.

(a) A number of designs used currently in the research lab of the author are shown here,

including multichannel input designs (which enable the fluid environment of a biological

sample in the central sample chamber to be exchanged rapidly in less than 1 s), microwells

(which have no fluid flow, but consists of a simple PDMS mask placed over living cells on a

microscope coverslip, here shown with bacteria, which can be used to monitor the growth of

separate cell “microecologies”), a wedge design that uses fluid flow to push single yeast cells

into the gaps between wedges in the PDMS design and in doing so immobilize them and thus

enable them to be monitored continuously using light microscopy with the advantage of not

requiring potentially toxic chemical conjugation methods, and a jail-​type design that consists

of chambers of yeast cells with a PDMS “lid” (which can be opened and closed by changing the

fluid pressure in the flow cell, which enables the same group of dividing cells to be observed by

up to eight different generations and thus facilitates investigation of memory effects across cell

generations). (b) A simple testing rig for bespoke microfluidics designs, as illustrated from one

used in the author’s lab, can consist of a simple gravity-​feed system using mounted syringes,

combined with a standard “dissection” light microscope that allows low magnification of a factor

of ca. 10–​100 to be used on the flow cell to monitor the flow of dyes or large bead markers.