Key Laboratory for Ultrafine Materials of Ministry of Education Centre for Biomedical Technologies Current tissue engineering technology aims to achieve the regeneration of human tissues, which integrates the key factors such as scaffolds, cells and biomolecules. Among these key factors, the development of high-performance scaffolds is the basis for the success of tissue engineering strategies. In the past decades, hydrogel scaffolds have been developed rapidly and widely used in biomedical field, however, their drawbacks have also been revealed, which shows that a single hydrogel scaffold cannot meet the excellent performance required in the field of tissue engineering. Recently, microspheres have been further engineered to fabricate structurally and functionally reliable artificial three-dimensional scaffolds of desired shape with enhanced specific biological functions. Therefore, the effective combination of hydrogel and microspheres can facilitate the development of high-performance scaffolds for tissue engineering and further fine-tuning the composite structure, which is expected to solve the dilemma faced by a single scaffold. In this review paper, we systematically summurized the type and preparation method for synthesis of hydrogel and microsphere materials commonly used in developing microsphere-containing hydrogel scaffolds. We then reviewed the broad application of these hybrid scaffolds in various fields of tissue engineering, followed by a summary and perspective on future directions.