Fungal species perform many important roles in biotechnology and recycling and act as agents of disease and decay. Surface-enhanced Raman scattering (SERS) has attracted significant attention as an analytical method for chemical and biological identification. For SERS experiments, it is essential to generate gold nanoparticles (AuNPs) with proper sizes and shapes. Raman and SERS imaging of fungi via in vivo synthesis of AuNPs were used to explore cellular components of Aspergillus nidulans (A. nidulans) cell. Critical parameters including pH, temperature and metal concentration affect the sizes and shapes of the NPs. For better control of NP formation (size, shape and location), pre-formed NP were incubated with A. nidulans colonies. Aspergillus nidulans outer hyphal walls were coated with NPs. Raman and SERS imaging of fungal walls revealed that proteins, carbohydrates and lipids are the main constituents of fungal cell wall.