This study aimed to investigate the effects of microbes on the composition of archaeological sandstone at the Temple of Isis in Qus city, Egypt. Six samples of degraded sandstone were collected to evaluate the deterioration process and analyzed by using X-Ray diffraction (XRD), Scanning electron microscopy (SEM), and USB digital microscope. Microbial isolation revealed the presence of microorganisms such as Aspergillus niger, Penicillium sp, Fusarium sp, and Bacillus sp. These microorganisms play a main role in the degradation of sandstone. X-ray diffraction analysis identified minerals like quartz, calcite, anhydrite, and gypsum, influencing its susceptibility to microbial deterioration. Scanning electron microscopy (SEM) showed the microbial populations, microstructure, and the formation of biofilms and erosion on the sandstone. The USB digital microscope examination provided detailed insights into the extent of degradation, including pigmented spots, micro cracks, and granular disintegration. The research highlights the complex interactions between microorganisms and the sandstone substrate, contributing to the understanding of biodeterioration in cultural heritage preservation.