Lipid implants have been proposed as promising sustained release devices for the parenteral application of pharmaceutical proteins. Near infrared spectroscopy (NIRS) has been reported in the literature to be a non-destructive tool for drug quantification within controlled release matrix systems based on poly-(lactic-co-glycolic) acid (PLGA). The objective of this study was to evaluate the potential application of NIRS for protein content determination within lipid matrices containing stabilizing and release modifying additives. Bovine serum albumin (BSA) and rh-interferon α-2a (IFN α-2a) were initially lyophilized with trehalose and then blended with tristearin (matrix material) and optionally with polyethylene glygol 6000 (PEG, release modifier). Implants were prepared by compression. NIR transmittance spectra were measured on a NIRTab® spectrometer. Partial least squares regression (PLSR) calibration models were developed to predict protein content in implants from the NIRS results. Additional samples were measured after performing release studies. It could be shown that NIRS allowed protein quantification in complex matrix systems with good accuracy after implant manufacture and during release studies [e.g., standard error of prediction (SEP) between 57 μg–176 μg]. In addition, small protein amounts down to 70 μg of incorporated protein per implant could be determined, thus demonstrating the low detection limit of NIRS.