An object of the present invention is to provide a method of detecting a specific tissue or cell in a sample tissue section and accurately specifying both the position(s) and amount of a biological substance of interest that is expressed on the specific tissue or cell. The method of quantifying a biological substance in a tissue section according to the present invention comprises: (1) performing bright-field observable immunostaining that specifically stains a first biological substance in the tissue section (first immunostaining); (2) performing immunostaining with a fluorescent substance-containing nanoparticle that specifically stains a second biological substance in the tissue section (second immunostaining); (3) specifying the expression position(s) of the second biological substance in the tissue section by comparing the position of a stained image of the first immunostaining and the position of a stained image of the second immunostaining; and (4) determining the expression amount of the second biolog
