Entrapment of fluorescence signaling DNA enzymes in sol-gel-derived materials for metal ion sensing.
Three fluorescence signaling DNA enzymes (deoxyribozymes or DNAzymes) were successfully immobilized within a series of sol-gel-derived matrixes and used for sensing of various metal ions. The DNAzymes are designed such that binding of appropriate metal ions induces the formation of a catalytic site that cleaves a ribonucleotide linkage within a DNA substrate. A fluorophore (fluorescein) and a quencher (DABCYL, [4-(4-dimethylaminophenylazo)benzoic acid]) were placed on the two deoxythymidines flanking the ribonucleotide to allow the generation of fluorescence upon the catalytic cleavage at the RNA linkage. In general, all DNAzymes retained at least partial catalytic function when entrapped in either hydrophilic or hydrophobic silica-based materials, but displayed slower response times and lower overall signal changes relative to solution. Interestingly, it was determined that maximum sensitivity toward metal ions was obtained when DNAzymes were entrapped into composite materials containing approximately 40% of methyltrimethoxysilane (MTMS) and approximately 60% tetramethoxysilane (TMOS). Highly polar materials derived from sodium silicate, diglycerylsilane, or TMOS had relatively low signal enhancements, while materials with very high levels of MTMS showed significant leaching and low signal enhancements. Entrapment into the hybrid silica material also reduced signal interferences that were related to metal-induced quenching; such interferences were a significant problem for solution-based assays and for polar materials. Extension of the solid-phase DNAzyme assay toward a multiplexed assay format for metal detection is demonstrated, and shows that sol-gel technology can provide new opportunities for the development of DNAzyme-based biosensors.
Shen
Yutu
Y
Department of Chemistry, McMaster University, Hamilton, Ontario, Canada, L8S 4M1.
Mackey
Gillian
G
Rupcich
Nicholas
N
Gloster
Darin
D
Chiuman
William
W
Li
Yingfu
Y
Brennan
John D
JD
D015374:Biosensing Techniques / Q000379:methods*; D002384:Catalysis; D021881:DNA, Catalytic / Q000737:chemistry*; D019793:Fluorescein / Q000737:chemistry; D005453:Fluorescence; D005782:Gels / Q000737:chemistry; D007477:Ions / Q000032:analysis; D008670:Metals / Q000032:analysis*; D012680:Sensitivity and Specificity; D012822:Silicon Dioxide / Q000737:chemistry; D013050:Spectrometry, Fluorescence / Q000379:methods; D013499:Surface Properties; D013997:Time Factors; D004124:p-Dimethylaminoazobenzene / Q000031:analogs & derivatives / Q000737:chemistry
D021881:DNA, Catalytic; D005782:Gels; D007477:Ions; D008670:Metals; C063127:4-(4-dimethylaminophenylazo)benzoic acid; D012822:Silicon Dioxide; D004124:p-Dimethylaminoazobenzene; D019793:Fluorescein