This review discusses label-free cell separation and sorting techniques in microfluidic systems, highlighting their importance in various biomedical applications such as diagnostics, therapeutics, and cell biology research. The authors emphasize the advantages of microfluidic methods, including automation, reduced cost, and the ability to handle single cells. They analyze various microfluidic techniques, focusing on their separation mechanisms, performance metrics, and recent developments. Key techniques include microscale filters, hydrodynamic filtration, deterministic lateral displacement (DLD), field-flow fractionation (FFF), microstructure-based separation, inertial microfluidics, gravitational and sedimentation methods, biomimetic approaches, magnetophoresis, aqueous two-phase systems (ATPS), acoustophoresis, and dielectrophoresis. Each technique is evaluated based on its separation criteria, such as size, shape, density, and electrical properties, and its potential for practical applications. The review also addresses the challenges and limitations of these methods, providing insights into their current state and future directions.This review discusses label-free cell separation and sorting techniques in microfluidic systems, highlighting their importance in various biomedical applications such as diagnostics, therapeutics, and cell biology research. The authors emphasize the advantages of microfluidic methods, including automation, reduced cost, and the ability to handle single cells. They analyze various microfluidic techniques, focusing on their separation mechanisms, performance metrics, and recent developments. Key techniques include microscale filters, hydrodynamic filtration, deterministic lateral displacement (DLD), field-flow fractionation (FFF), microstructure-based separation, inertial microfluidics, gravitational and sedimentation methods, biomimetic approaches, magnetophoresis, aqueous two-phase systems (ATPS), acoustophoresis, and dielectrophoresis. Each technique is evaluated based on its separation criteria, such as size, shape, density, and electrical properties, and its potential for practical applications. The review also addresses the challenges and limitations of these methods, providing insights into their current state and future directions.