Biomedical Engineering Reference
In-Depth Information
be immobilized onto micromatrix gel pads containing aldehyde groups [25,27].
Oligonucleotides have also been immobilized on glass slides coated with an ac-
tivated agarose film [28]; the outcome of hybridizations with longer labelled
fragments was less reliable on these slides than on conventional aldehyde-
functionalized glass slides.
3.2.5 Fiber Optic Arrays
This technique utilizes probes that are immobilized onto microspheres us-
ing well established procedures [29-32]. Probe-functionalized microspheres
are coupled to high density fiber optic arrays; the optical fiber substrate al-
lows simultaneous and repetitive monitoring of the microsphere array [30,33].
Amine-modified oligonucleotides are activated by treatment with cyanuric
chloride, and then reacted with polyethyleneimine (PEI)-coated microspheres.
The beads are then rinsed with a sodium borate buffer, and the unreacted
amine groups on the beads are capped using succinic anhydride to prevent
non-specific binding of DNA.
3.2.6 Polymers
Polymers like nylon and polypropylene have been used for arraying oligonu-
cleotides. Oligonucleotides have been immobilized onto nylon supports us-
ing UV crosslinkers [34-36]. High density arrays have been constructed on
aminated polypropylene supports using phosphoramidite chemistry [37-39].
Oligonucleotides have been immobilized onto polypropylene supports co-
valently, using bifunctional crosslinkers or EDC-mediated amide bond for-
mation between amine-terminated oligonucleotide and carboxylate-modified
polypropylene plates [40, 41], and non covalently [42]. Non-covalently im-
mobilized oligonucleotides are, however, susceptible to removal under high
salt/high temperature conditions.
3.3 Protein Microarrays
The commercial development of protein microarrays has been di
cult in great
part due to the increased complexity that comes with dealing with proteins
(compared to oligonucleotides or cDNA). Proteins tend to denature on sur-
faces [43, 44]; this denaturation can result in a loss of their activity. Proteins
also tend to adsorb non-specifically on a wide variety of surfaces [45]; this non-
specific adsorption can lead to the misinterpretation of the results of microar-
ray experiments. On account of these challenges, surface functionalization and
protein immobilization procedures are very important for the successful im-
plementation of protein microarrays. A wide variety of substrates and surface
chemistries have been used in academic research and some have been devel-
oped commercially. Some of these methods are described below; Table 3.2 lists
several surfaces available commercially.
