4.20.7.3.2. The Affymetrix Gene Chip Array Technology

4.20.7.4. The Principle of Microarray Technology

The main principle behind microarray technology is the hybridization between two DNA strands. The property of complementary nucleic acid sequences to specifically pair with each other by forming hydrogen bonds between complementary nucleotide base pairs is the main force of recognizing complementary partner. A high number of complementary base pairs in a nucleotide sequence allow tighter non-covalent bonding between the two strands. After washing off of non-specific bonding sequences, only strongly paired strands will remain hybridized. So fluorescently labeled target sequences that bind to a probe sequence generate a signal that depends on the strength of the hybridization determined by the number of paired bases, the hybridization conditions such as temperature, and washing after hybridization. Total strength of the signal, from a spot, depends upon the amount of target sample binding to the probes present on that spot. Microarrays use relative quantification in which the intensity of a spot is compared to the intensity of the same spot under a different condition, and the identity of the spot is known by its position. The figure below explains the steps involved in a microarray.

Figure 4.52: The major step involved in a microarray experiment