1：Experimental Design and Method Selection:

The study involved synthesizing HSA-stabilized copper nanoclusters (HSA-CuNCs) at room temperature using hydrazine hydrate as a reducing agent and HSA as a stabilizing template. The fluorometric method was chosen for bilirubin detection due to its high selectivity, sensitivity, and cost-effectiveness.

2：Sample Selection and Data Sources:

Human serum albumin, bilirubin, and other chemicals were purchased from suppliers. Human blood serum and urine samples were obtained from a hospital for real sample analysis.

3：List of Experimental Equipment and Materials:

Instruments included JASCO FP-6600 spectrofluorometer, JASCO V-630 UV-visible spectrophotometer, JEOL JEM 2100 HR-TEM, Horiba Scientific TCSPC unit, Shimadzu FT-IR 8000 spectrophotometer, Zeta-sizer Nano ZS, and Bruker Advanced D8 XRD. Materials included HSA, CuSO4, N2H

4：H2O, NaOH, and various interfering substances. Experimental Procedures and Operational Workflow:

HSA-CuNCs were synthesized by mixing HSA with CuSO4, adjusting pH to ~12 with NaOH, adding N2H

5：H2O, stirring for 240 min, and purifying by dialysis. Bilirubin detection involved mixing bilirubin with HSA-CuNCs in PBS buffer, equilibrating for 6 min, and measuring emission spectra at 390 nm excitation. Data Analysis Methods:

Emission intensity changes were analyzed using Stern-Volmer and double logarithmic equations to determine quenching constants and binding affinity. Statistical analysis included calculating limits of detection and recovery percentages.