Vibrio parahaemolyticus (V. parahaemolyticus) is an important foodborne pathogen known to cause severe enteric disease. Thus, timely detection of V. parahaemolyticus in seafood is crucial to prevent food poisoning and reduce economic losses. Traditional lateral flow immunoassay strips (LFIS) required good labelling materials and pairing of two antibodies, which made them costly and difficult to manufacture. In this study, a label-free and lac dye coloration-based LFIS (LD-LFIS) to detect trh + V. parahaemolyticus was developed for the first time. Lac dye was used to stain V. parahaemolyticus, and LFIS was used to detect stained bacteria. Dimethyl sulphoxide (DMSO) and simultaneous mordanting were chosen as the best solvent and the best staining method for lac dye. In addition, three mordants [KAl(SO 4 ) 2 ·12H 2 O, NH 4 Fe(SO 4 ) 2 ·12H 2 O, and AlCl 3 ·6H 2 O] were selected to improve dyeing efficiency. The detection limit of LD-LFIS was 3.9 × 10 5 CFU/ml when NH 4 Fe(SO 4 ) 2 ·12H 2 O was used as mordant. Feasibility of the LD-LFIS method was verified by detecting trh + V. parahaemolyticus in true and spiked seafood samples. The method developed in this study is expected to reduce restrictions on labelling materials and pairing of two antibodies on LFIS, and proposes a novel idea for the rapid detection of V. parahaemolyticus in seafood.