Table 3 Characterization methods for primary osteoblasts in dental research [9, 13, 20, 29,30,31,32,33,34,35,36,37,38]
Technique | Purpose | Markers/assays used | Advantages | Limitations |
|---|---|---|---|---|
Morphological analysis | Confirms characteristic osteoblastic shape | Phase-contrast microscopy, light microscopy | Simple, fast, non-invasive | Insufficient alone to confirm identity |
ALP assay | Evaluates early differentiation and activity | Spectrophotometric ALP enzyme activity | Indicates functional osteoblast activity | Limited to early differentiation |
Gene expression analysis | Confirms osteoblast phenotype at molecular level | qPCR, RT-PCR (Runx2, ALP, COL1A1, OCN) | Precise molecular insights | Requires RNA extraction and careful handling |
Immunocytochemistry/Western blotting | Validates protein expression of osteoblast markers | Osteocalcin, osteopontin, COL1 | Confirms phenotype at protein level | Labor-intensive, requires specific antibodies |
Mineralization assays | Assesses ability to form mineralized matrix | Alizarin Red, von Kossa staining | Visual confirmation of calcium and phosphate deposition | Semi-quantitative, may require complementary assays |
SEM | Examines surface morphology and ECM organization | High-resolution SEM imaging | Evaluates cell–material interactions | Requires expensive equipment and preparation |
Proliferation assays | Measures metabolic activity and growth | MTT, BrdU, XTT assays | Quantifies cell viability and biocompatibility | Cannot differentiate osteoblasts from other cell types |