The human body harbors diverse microbial communities essential for maintaining health and influencing disease processes. Droplet microfluidics, a precise and high-throughput platform for manipulating microscale droplets, has become vital in advancing microbiome research. This review introduces the foundational principles of droplet microfluidics, its operational capabilities, and wide-ranging applications. We emphasize its role in enhancing single-cell sequencing technologies, particularly genome and RNA sequencing, transforming our understanding of microbial diversity, gene expression, and community dynamics. We explore its critical function in isolating and cultivating traditionally unculturable microbes and investigating microbial activity and interactions, facilitating deeper insight into community behavior and metabolic functions. Lastly, we highlight its broader applications in microbial analysis and its potential to revolutionize human health research by driving innovations in diagnostics, therapeutic development, and personalized medicine. This review provides a comprehensive overview of droplet microfluidics' impact on microbiome research, underscoring its potential to transform our understanding of microbial dynamics and their relevance to health and disease.