Surface-enhanced Raman spectroscopy (SERS) substrates of silver nanoflowers were prepared based on lyotropic liquid crystals for the quantitative detection of neoCoronavirus spike protein (S protein). Firstly, a Raman spectroscopy detection scheme for neoCoronavirus was designed, and the SERS substrate was prepared using lyotropic liquid crystal soft templates and electrochemical deposition techniques. Then, spectral consistency and inter-substrate detection differences were evaluated to ensure reliable quantitative detection. Then, the sensitivity of the SERS substrate for the detection of S protein was analysed, and its detection limit was 2×10^-6 mg/mL. Finally, the quantitative detection method was established, yielding a precise concentration-intensity relationship with a measurement error of 4×10^-7 mg/mL. In this paper, the technique of preparing SERS substrates based on lyotropic liquid crystals was used for the high sensitivity detection of neoCoronavirus spike protein, and also provides a feasible technical way for the detection of other proteins.