Metasurfaces exhibit excellent optical performance to enhance the light–matter interaction of target molecules in biosensing based on its well‐optimized nanostructured unit cells. In the meantime, fluorescence (FL) biosensors with aggregation induced emissions (AIE) features also demonstrate outstanding performance in biomarker detection due to their fast response, high selectivity, and low background noise. Nevertheless, extremely low‐level analytes are difficult to detect in practical applications since complex urine samples include a number of uncontrolled variables such as impurities, autofluorescence, other urine components, etc. At present, improving optical signal sensitivity of human serum albumin (HSA) detection is always a big challenge to overcome such interference in human urine scenarios. In this work, first an AIE‐based FL biosensor TPE‐4TA is combined with an all‐dielectric metasurface platform to achieve quantitative detection of trace HSA in urine by utilizing biofunctionalization protocols on the silicon (Si) nanostructures. The results indicate significant FL enhancement in the metasurface platform that offers a promising pathway for improving biomarker detection in the future.