The confluent HA-VSMCs were washed with PBS twice then the cell layers were scraped into a solution. The cell lysates were homogenized and measured for ALP activity by ALP assay kit (A059-2, Jiancheng, Nanjing) and for osteocalcin secretory by osteocalcin assay kit (H152, Jiancheng, Nanjing) according to the manufacturer’s instructions. ALP activity and osteocalcin were normalized to total cellular protein of the cell layers by the Bradford protein assay as previously described [39 (link)].
Osteocalcin assay kit
Manufactured by Nanjing Jiancheng
Sourced in China
The Osteocalcin assay kit is a laboratory product designed to measure the levels of osteocalcin, a protein found in bone. Osteocalcin is an important marker for evaluating bone metabolism and health. The kit provides the necessary reagents and protocols to quantify osteocalcin concentrations in biological samples.
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2 protocols using osteocalcin assay kit
1
Measuring ALP Activity and Osteocalcin Secretion
2
Osteogenic Differentiation of VSMC
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HA-VSMC cells or transfected HA-VSMC cells were cultured with 30 mmol/L glucose for 14 days. Then, cells were washed, scraped into a solution, and lysed with Cell lysis buffer for Western and IP without inhibitors (Beyotime Biotechnology, Shanghai, China). Cell lysates were measured for alkaline phosphatase (ALP) activity using an Alkaline Phosphatase Assay Kit (Beyotime Biotechnology, Shanghai, China) and for osteocalcin secretion using an osteocalcin assay kit (Jiancheng, Nanjing). ALP activity and osteocalcin secretion were normalized to the total cellular protein of the cellular layers and then normalized to that of the control [21 (link)].
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