Gold nanoparticles (AuNPs) have drawn significant interest in recent years due to unique properties that make them advantageous in biomedical applications, including drug delivery and tissue engineering. In this paper, we have developed multiple methods for the synthesis of AuNPs using English ivy as the substrate. In the first method, we have used actively growing English ivy shoots to develop a sustainable system for the production of ivy nanoparticles. The second method was developed using the extract from the adventitious roots of English ivy. The nanoparticles formed using both methods were compared to determine the size distribution, morphology, and chemical structure of the nanoparticles. Characterization of the AuNPs was conducted using ultraviolet-visible (UV-Vis) spectroscopy, dynamic light scattering (DLS), scanning electron microscopy (SEM), and energy-dispersive X-ray spectroscopy (EDS). In addition to the structural differences between the AuNPs formed from the different methods, details of the methods in terms of yield, duration, and speed of AuNP formation are also discussed. Further, this paper will show that AuNPs formed using both methods demonstrated efficient uptake in mammalian cells, which provides the potential for biomedical applications. The two methods developed through this research for eco-friendly synthesis of AuNPs present an alternative to traditional chemical synthesis methods.