In this study, the effect of Achyranthes bidentata alcohol on the proliferation of osteoblasts was detected via methyl thiazolyl tetrazolium (MTT) assay. 
MTT
The effect of Achyranthes bidentata alcohol on the alkaline phosphatase (ALP) activity in osteoblasts was analyzed via ALP assay. 
ALP assay
The effect of Achyranthes bidentata alcohol on the expression of osteoblast marker gene, Runt-related transcription factor 2 (Runx2), was detected via reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and immunohistochemistry. 
RT-qPCR
Moreover, the phosphorylation or activation of extracellular signal-regulated kinase (ERK) in osteoblasts induced by Achyranthes bidentata alcohol was analyzed using western blotting. 
western blotting