Scanning Electrochemical Microscopy Imaging of DNA Arrays for High Throughput Analysis Applications - Applications of Electrochemistry and Nanotechnology in Biology and Medicine - page 108

1. Types and Manufacture Methods of DNA Arrays
The earliest DNA arrays may be classed as filter arrays that were
able to accommodate the Southern blot technique and the many
derivatives thereof. Subsequent improvements led to the creation
of high-density filter arrays, termed macroarrays. Subsequently,
two different microarray-based technologies arose: the DNA mi-
croarray, better known as the spotted microarray, developed at
Stanford University, 3 and the oligonucleotide arrays synthesized in
situ , such as Affymetrix GeneChips ® . 4
In preparing a DNA array, gene-specific probes are created
and immobilized on a solid support. When the DNA sequence in-
formation is available, oligonucleotides that hybridize with each
gene can be synthesized. This approach precludes the need to
manage large clone libraries as it is guided by sequence infor-
mation. Moreover, this strategy is particularly well suited to the
expression profile analysis of organisms with completely se-
quenced genomes, as the focused custom array can analyze all
predicted genes in a single experiment.
In the case of the DNA microarray, the method starts by syn-
thesizing cDNA from a cell's messenger RNA using reverse tran-
scriptase polymerase chain reaction. Robotically operated, small
pipettes or a piezoelectric device similar to an inkjet printer pro-
duces the microarrays by accurately placing large numbers of spots
of the cDNA onto a substrate, usually glass ( Fig. 1 ). This type of
DNA array can either be homemade on a microscope glass slide,
or custom made with 1,000 to 10,000 different spots per slide. Spot
sizes of 20 to 50 μm are commonly used and each spot can be an-
alyzed with a fluorometry scanner with a conventional design. This
type of DNA array is particularly suitable for in-house fabrication
of each experiment with its relative ease of custom array prepara-
tion by choosing the probes, the printing and the array settings.
The light-directed, combinatorial solid-phase chemistry
method can directly synthesize complete sets of oligonucleotide
probes on a substrate, as developed by Affymetrix Corp ( Fig. 2 ). In
this method, a fused silica substrate is coated with a hydroxyl ter-
minated, silane coupling agent to render the surface active. The
treatment is followed by the reaction with the DMT-hexa-ethyl-