Monitoring cell secretion in complex microenvironments is crucial for understanding cellular behavior and advancing physi-

ological and pathological research. While traditional cell culture methods, including organoids and spheroids, provide valuable

models, real-time monitoring of cell secretion of signaling molecules remains challenging. Integrating advanced monitoring

technologies into these systems often disrupts the delicate balance of the microenvironment, making it difficult to achieve sensitiv-

ity and specificity. This review explored recent strategies for integrating the monitoring of cell secretion of signaling molecules,

crucial for understanding and replicating cell microenvironments, within cell culture platforms, addressing challenges such as

non-adherent cell models and the focus on single-cell methodologies. We highlight advancements in biosensors, microfluidics,

and three-dimensional culture methods, and discuss their potential to enhance real-time, multiplexed cell monitoring. By examin-

ing the advantages, limitations, and future prospects of these technologies, we aim to contribute to the development of integrated

systems that facilitate comprehensive cell monitoring, ultimately advancing biological research and pharmaceutical development.