Human osteocalcin sandwich elisa kit

MSC-seeded scaffolds were harvested after 1, 7, 14 and 28 days of cultivation, washed with ice-cold PBS and individually homogenised in RIPA lysis buffer containing Halt TM protease inhibitor cocktails (both solutions from Thermo Scientific, UK). Three freeze-thaw cycles were then performed to promote cell lysis. Enzymatic activity of alkaline phosphatase (ALP) was detected using the clear supernatants from cell lysates and an ALP fluorometric assay kit (Source Bioscience, UK). Osteocalcin production was analysed using a human osteocalcin sandwich ELISA kit (Invitrogen) and following the manufacturer's protocol.
All samples and standards were done in triplicate. Cell-free scaffold samples were used as negative controls.

MSC-seeded scaffolds, after cultivation for 1, 7, 14 and 28 days in osteogenic medium, were harvested, washed with ice-cold PBS, homogenised in RIPA lysis buffer containing Halt TM protease inhibitor cocktails (Thermo Fisher Scientific, UK) and then freeze-thawed for 3 cycles to promote cell lysis. Alkaline phosphatase (ALP) activity was detected using the clear supernatants from cell lysates and an ALP fluorometric assay kit (Source Bioscience, UK).
Osteocalcin production was analysed using a human osteocalcin sandwich ELISA kit (Invitrogen) and following the manufacturer's protocol. All samples and standards were done in triplicate. Cell-free scaffold samples were used as negative controls.