A method for highly sensitive determination of a target substance by means of an aptamer includes: causing an aptamer capable of specific binding with a target substance to bind competitively with said target substance and a nucleic acid strand having a base sequence complementary to at least a portion of said target substance, detecting at least either of the physical change and the chemical change that results from said aptamer binding with said nucleic acid strand, and determining said target substance based on the result of detection.