Abstract: Starter cultures are regarded as the “chips” of fermented foods, playing a pivotal role in enhancing sensory attributes, nutritional value, probiotic functionality, and production efficiency. Therefore, the exploration and preparation of starter cultures are of great significance for the high-quality development of the fermentation industry. This review summarizes recent advances in modern starter culture preparation technologies, with emphasis on strain screening, high density culture, and freeze-drying. Efficient screening strategies, such as automation, microfluidics, and artificial intelligence, are highlighted for improving accuracy and throughput in microbial strain screening. The limitations and optimization strategies of high-density culture are systematically analyzed, with a focus on fed-batch and multi-stage continuous culture to improve biomass yield. Furthermore, the mechanisms of microbial inactivation during freeze-drying and corresponding protection strategies, such as optimization of culture conditions, stress adaptation, addition of protective agents, and genetic engineering, are elaborated to enhance cell viability and stability. This review aims to provide theoretical insights and practical guidance for the efficient exploration of food microorganisms and the development of high-density and stable starter cultures.