Black Soldier Fly (Hermetia illucens) larvae have emerged as an effective bioconversion agent capable of transforming organic waste into high-protein biomass suitable for animal feed applications. This study aimed to analyze the effectiveness of liquid milk waste feeding on the protein content of Black Soldier Fly larvae by optimizing feed amount and harvest age using Response Surface Methodology (RSM). A laboratory-scale experiment was conducted using Central Composite Design (CCD) with 13 experimental runs. Four-day-old Black Soldier Fly larvae (30 g per treatment) were fed varying amounts of liquid milk waste (2.3–3.7 liters) and harvested at different ages (4–10 days). Environmental conditions were monitored daily, maintaining temperatures of 28.8–29.7°C and humidity of 71–75%. Protein content was determined through laboratory analysis, and data were analyzed using ANOVA and RSM optimization. The highest protein content achieved was 12.93% at 3.5 liters feed amount with harvest on day 5, while the lowest was 10.18% at 3 liters feed with harvest on day 10. The quadratic model demonstrated excellent fit (R² = 0.9666), with both feed amount (p = 0.0001) and harvest age (p < 0.0001) significantly affecting protein content. RSM optimization suggested optimal conditions of 3.4 liters feed with day-5 harvest, predicting 13.034% protein content. Validation experiments confirmed this prediction with 12.96% protein content. In Conclusion, Liquid milk waste effectively supports Black Soldier Fly larvae protein production, demonstrating the potential for sustainable waste valorization and alternative protein source development.