In the context of increasingly airtight homes, there is currently little known about the type and diversity of microorganisms in the home, or factors that could affect their abundance, diversity and nature. In this study, we examined the type and prevalence of cultivable microorganisms at eight different sites in 100 homes of older adults located in Glasgow, Scotland. The microbiological sampling was undertaken alongside a household survey that collated information on household demographics, occupant behaviour, building characteristics, antibiotic use and general health information. Each of the sampled sites revealed its own distinct microbiological character, in both species and number of cultivable microbes. While some potential human pathogens were identified, none were found to be multidrug resistant. We examined whether the variability in bacterial communities could be attributed to differences in building characteristics, occupant behaviour or household factors. Sampled sites furnished specific microbiological characteristics which reflected room function and touch frequency. We found that homes that reported opening windows more often were strongly associated with lower numbers of Gram-negative organisms at indoor sites (p < 0.0001). This work offers one of the first detailed analysis of cultivable microbes in homes of older adults and their relationship with building and occupancy related factors, in a UK context.