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We present a novel method for on-chip competitive dispersion immunoassay (DISIA) which aims at measuring the specificity between fluorescently-labeled antibodies (Ab*) and label-free antigens (Ag). We have fabricated a nanofluidic device that allows fast detection of interaction of antibodies and antigens (< 30s) based on lateral dispersion of fluorescent species in a nanoslit, using small amounts of solution (< 1μl). We measured that doubling the weight of streptavidin protein results in an increase of 30% of the lateral dispersion of fluorescent coupled molecules in the nanoslit, which can be read easily.