Optimization of preparation for oligonucleotide array and hybridization

Silanization of clean glass slides with 3% AFTES (3'-aminopropyltrimethoxysilane) or 3% DETA (trimethoxysi-lylpropyldiethylenetriamine) was performed by incubation with different time respectively, followed by disposal with 5% glutaralde-hyde in water for 1 hour. The results showed that incubation with 3% AFTES (dissolved by 95% acetone) for 2 hours was enough to get strong fluorescence signal. Compared with polyT15 spacer, probes with PEG6 and PEG12 spacer could improve signal intensity to 6-and 8-folds. When using carbonate buffer with 9-11 pH value as spotting solution, there's no significant decrease of signal intensities as pH value grows up. Best signal intensity could be produced when 25μM oligonucleotide probe hybridized with 2μL fluorescence labeled 110bp asymmetry PCR product which was diluted to 10 μL by 6 × SSEPT buffer.