An inspecting method for measuring, using a test substance-containing solution containing a test substance and a liquid, a concentration of the test substance, which is contained in the test substance-containing solution, in the liquid, the inspecting method using an inspecting instrument (1, 11), the inspecting instrument (1, 11) including a wall (3, 13, 33) that has a periodic structure (3A, 13A, 33A) resulting from a plurality of recesses (3a, 13a, 33a) or a plurality of protrusions, the plurality of recesses (3a, 13a, 33a) or the plurality of protrusions including a refractive index adjusting layer (7, 17, 37) on surfaces thereof, the refractive index adjusting layer (7, 17, 37) being a silicon layer, the inspecting method comprising the steps of: measuring a reference light intensity by irradiating the periodic structure with light while the liquid has been introduced into the inspecting instrument; measuring an evaluation light intensity by irradiating the periodic structure with light while the test substance-containing solution has been introduced into the inspecting instrument and the test substance and the liquid are present; and acquiring a relative value between the reference light intensity and the evaluation light intensity, and the inspecting method comparing the relative value obtained when a specimen with a known concentration of the test substance is introduced into the inspecting instrument, to the relative value obtained when a specimen with an unknown concentration of the test substance is introduced into the inspecting instrument, to determine the concentration of the test substance in the specimen with the unknown concentration of the test substance. the inspecting method using an inspecting instrument (1, 11), the inspecting instrument (1, 11) including a wall (3, 13, 33) that has a periodic structure (3A, 13A, 33A) resulting from a plurality of recesses (3a, 13a, 33a) or a plurality of protrusions, the plurality of recesses (3a, 13a, 33a) or the plurality of protrusions including a refractive index adjusting layer (7, 17, 37) on surfaces thereof, the refractive index adjusting layer (7, 17, 37) being a silicon layer, the inspecting method comprising the steps of: measuring a reference light intensity by irradiating the periodic structure with light while the liquid has been introduced into the inspecting instrument; measuring an evaluation light intensity by irradiating the periodic structure with light while the test substance-containing solution has been introduced into the inspecting instrument and the test substance and the liquid are present; and acquiring a relative value between the reference light intensity and the evaluation light intensity, and the inspecting method comparing the relative value obtained when a specimen with a known concentration of the test substance is introduced into the inspecting instrument, to the relative value obtained when a specimen with an unknown concentration of the test substance is introduced into the inspecting instrument, to determine the concentration of the test substance in the specimen with the unknown concentration of the test substance. measuring a reference light intensity by irradiating the periodic structure with light while the liquid has been introduced into the inspecting instrument; measuring an evaluation light intensity by irradiating the periodic structure with light while the test substance-containing solution has been introduced into the inspecting instrument and the test substance and the liquid are present; and acquiring a relative value between the reference light intensity and the evaluation light intensity, and the inspecting method comparing the relative value obtained when a specimen with a known concentration of the test substance is introduced into the inspecting instrument, to the relative value obtained when a specimen with an unknown concentration of the test substance is introduced into the inspecting instrument, to determine the concentration of the test substance in the specimen with the unknown concentration of the test substance.