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Osteoclast generation from RAW 264.7 and PBMC cells. The set up in our lab

Susana Jurado, Albert Parés, Pilar Peris, Andrés Combalia, Ana Monegal Brancos, Nuria Guañabens

Introduction and objectives: osteoclasts are terminally differentiated giant multinucleated cells derived from the fusion of mononuclear progenitors of the monocyte/macrophage hematopoietic lineage. The objective of our group was to achieve the best method for osteoclast differentiation, from both RAW 264.7 cells and peripheral blood monocytes. Material and methods: RAW 264.7 cells and human PBMCs were differentiated into osteoclasts. Success in differentiation was assessed by TRAP staining. Osteoclast activity was detected by the resorption pits in Corning® Osteo Assay Surface Plates. Results: the optimal cell density for RAW 264.7 cell differentiation was 25,000 cells/cm2 with 30 ng/mL of RANKL for 6 days. Osteoclasts differentiated from PBMCs were observed after 4 weeks with 25 ng/mL M-CSF and 30 ng/mL RANKL. Individual pits or multiple pit clusters were observed on the surface plates. Conclusions: we report optimal conditions for the differentiation of osteoclasts from RAW 264.7 and PBMCs cells to carry out our future experiments.

Palabras Clave: Osteoclasts. Differentiation. Bone resorption.

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