Here, we report a novel method for micropatterning oligonucleotides on the diamond surface via forming amine groups on the diamond surface by nitrogen/hydrogen radical treatment. The covalent bonding of the supporting oligonucleotide and characterization of an immobilized hybridized oligonucleotide with Cy5 modification were investigated by fluorescence microscopy. To investigate the effectiveness of nitrogen/hydrogen radical treatment for amine termination, two types of radical treatment were used: hydrogen/nitrogen radical treatment and pure nitrogen radical treatment. From the results, hydrogen/nitrogen radical treatment produces amine (NH2) termination on the diamond surface. The effect of amine termination was investigated by immobilization of single-stranded DNA via amide bonding between surface NH2 groups and COOH groups terminating the DNA. The immobilized single-stranded DNA (supporting DNA), which has a complementary relationship with the adenosine triphosphate (ATP) aptamer (DNA), hybridizes with the aptamer with attached fluorescence dye. When ATP molecules approach the double-stranded DNA, the aptamer forms a close relationship with the supporting DNA and combines with ATP. ATP detection was effectively carried out by reduction of fluorescence.