title: Creating highly dense and uniform protein and DNA microarrays through photolithography and plasma modification of glass substrates.
abstract: We demonstrate a method to create high density protein microarrays with excellent spot uniformity using photolithography and plasma processing on low cost commercially available microscope glass slides. Protein deposition and fluorescence signal evaluation on these substrates are performed by standard arrayers and scanners. To this end, spots of commercial photoresists (AZ5214, SU8 and Ormocomp(®)) were defined through lithography on glass substrates followed by short SF(6) plasma treatment and selective protein adsorption on these spots with respect to glass (spot to background fluorescence signal ratios 30:1 to 40:1) was demonstrated using model protein binding assays. Among the photoresists tested, Ormocomp was selected since it provided the highest protein binding capacity. No ageing of Ormocomp/glass substrates in terms of protein binding capacity was observed for at least two months. Besides to protein microarrays, DNA microarrays were also developed by spotting streptavidin-biotinylated oligonucleotide conjugates corresponding to wild- and mutant-type sequences of four deleterious BRCA1 gene mutations. For all of the examined mutations, higher specific hybridization signals (1.5-4 times) and improved discrimination ratios between wild- and mutant-type sequences as well as higher spot uniformity and repeatability were demonstrated on Ormocomp/glass substrates with intra- and inter-spot CVs of 8.0% and 4.5%, respectively, compared to commercial polystyrene (intra- and inter-spot CVs 36% and 18%) and epoxy-coated glass (intra- and inter-spot CVs 26% and 20%) slides. Thus, the proposed substrates can be readily applied to protein and DNA microarrays fabrication and, moreover, the described method for selective protein adsorption can be advantageously implemented in various analytical microdevices for multi-analyte detection.
authors.affiliation: Institute of Microelectronics, NCSR Demokritos, Attiki, Greece.
authors.forename: A
authors.initials: A
authors.lastname: Malainou
authors.forename: P S
authors.initials: PS
authors.lastname: Petrou
authors.forename: S E
authors.initials: SE
authors.lastname: Kakabakos
authors.forename: E
authors.initials: E
authors.lastname: Gogolides
authors.forename: A
authors.initials: A
authors.lastname: Tserepi
mesh_terms: D000327:Adsorption; D019313:BRCA1 Protein / Q000302:isolation & purification; D015374:Biosensing Techniques / Q000295:instrumentation / Q000379:methods*; D005453:Fluorescence; D005898:Glass / Q000737:chemistry; D006801:Humans; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation / Q000379:methods*; D010949:Plasma / Q000737:chemistry*; D040081:Protein Array Analysis / Q000295:instrumentation / Q000379:methods*; D011485:Protein Binding; D019809:Streptavidin / Q000737:chemistry; D013499:Surface Properties
chemical_list: D019313:BRCA1 Protein; C492913:BRCA1 protein, human; D019809:Streptavidin