The objective of the present study was to develop a rapid and sensitive HPLC-florescence detection method for the quantification of silodosin in pharmaceuticals using “the Analytical Quality by Design (AQbD)” approach. For this purpose, at first, spectrofluorometric measurments were conducted to determine optimum excitation and emission wavelengths for silodosin, and they were found as 226 and 456 nm, respectively. A central composite design methodology was applied for optimization of critical method parameters. The parameters that have an impact on chromatographic separation of silodosin were selected as pH, column temperature, and organic content of the mobile phase (acetonitrile %) considering previous studies in the literature. A quadratic three-factor central composite design model consisting of 20 experimental observations was used for optimization of the parameters. According to the response surface methodology, the optimized conditions for the column temperature, acetonitrile percentage and the pH of the mobile phase were found as 36.8 °C, 28.2% and 3.3, respectively. The optimized method was validated according to ICH guidelines for accuracy, precision, working range, reproducibility, the limit of detection, the limit of quantification, and robustness. The method was linear in the range of 0.1–40 µg/mL, with a high correlation coefficient (0.9991) and acceptable precision (RSD<7.8%). Using the AQbD approach has provided advantages in terms of time consumption and costs. After validation studies, the developed method was successfully applied in the analysis of silodosin-containing tablet formulation indicating that the method could be used for routine quality control analyses.