The invention discloses a water-soluble asymmetric indocyanine fluorescent dye. A general formula of the dye is as shown in the specification, wherein n is equal to 1, 2 or 3. A preparation method comprises the steps that (1), N-(3,5-2(2-(2-methoxyethoxy) oxethyl) oxethyl) benzyl-2,3,3-trimethyl-5-sulfonic group-3H-indole is synthesized; 2,3,3-trimethyl-5-sulfonic group-3H-indole, organic sylvine, and 3,5-2(2-(2-methoxyethoxy) oxethyl) oxethyl) oxethyl benzylbromide are added to an alcohol solvent in sequence, and conduct backflow reaction for 20-40h under inert gas shielding; the solvent is distilled off after the reaction; column separation is conducted; a red solid product is obtained; and (2), N-carboxyl benzyl-2,3,3-trimethyl-5-sulfonic group-3H-indole, the product and a condensing agent are dissolved in an acetic acid/acetic anhydride mixed solvent; pyridine or pyrimidine is added, and reacts at 60-100 DEG C for 1-6h under the inert gas shielding; ethyl acetate is added after the reaction for dissolving out a precipitate; suction filtration is conducted; the product is separated by taking a methanol/acetonitrile mixed solvent as an eluent, and using a reverse phase silica gel filling separation column; three color strips are obtained; the second color strip is accepted; and the water-soluble asymmetric indocyanine fluorescent dye is obtained.