Mass spectrometric characterization of DNA microarrays as a function of primary ion species

Recent studies have shown TOF-SIMS to be an appropriate method for the detailed examination of the immobilization process of PNA and its ability to hybridize to unlabeled complementary DNA fragments. Unlabeled single-stranded DNA was hybridized to Si wafer biosensor chips containing both complementary and non-complementary immobilized PNA sequences. The hybridization of complementary DNA could readily be identified by detecting phosphate-containing molecules from the DNA backbone. An unambiguous discrimination was achieved between complementary and non-complementary sequences. In order to optimize detection parameters, different primary ions were applied, including monoatomic ions (Bi+) as well as cluster ions (Bi2 +, Bi3 +, Bi4 +, Bi3 ++, Bi5 ++), and secondary ion yield behavior and formation efficiencies were studied. It was found that cluster primary ions resulted in a significantly increased yield of DNA-correlated fragments, enabling higher signal intensities and better secondary ion efficiencies. TOF-SIMS is undoubtedly a highly useful technique for identifying hybridized DNA on PNA biosensor chips. It is suitable for studying the complexity of the immobilization and hybridization processes and may provide a rapid method for DNA diagnostics. With the absence of the labeling procedure and the simultaneous increase of the phosphate signal as a result of increasing DNA sequence length, this technique comes to be especially useful for the direct analysis of genomic DNA.