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In Vitro Osteogenesis Assay Kit
Description:
In Vitro Osteogenesis Assay Kit
Trade Name:
Chemicon (Millipore)
Qty/Pk:
1 kit
Product Overview:
Introduction:
Bone matrix is a dense mixture comprised mainly of collagen fibers and calcium phosphate particles, with a population of living cells contained within. Despite its rigidity, bone continually undergoes a remodeling process that requires the coordinated activity of two types of cells, osteoclasts and osteoblasts [1]. Many diseases of bone including osteoporosis, a common phenomenon in post-menopausal women in which bone mass is greatly reduced, and osteogenesis imperfecta, also known as brittle-bone disease, are likely caused by the misregulation of osteoblasts and osteoclasts. Understanding the molecular mechanisms that underlie osteogenesis, the process by which new bone is formed, is thus of significant clinical importance [2].
Osteoclasts are related to macrophages, and erode the bone matrix by secreting acids and hydrolases to dissolve bone minerals and digest organic components. Osteoblasts arise from undifferentiated precursor cells, and deposit a mineralized matrix consisting of collagen, calcium, and phosphorous and other minerals, leading to the formation of new bone. Osteoblasts have also been shown to express factors which regulate the differentiation and function of osteoclasts [2]. The balance between the activity of osteoblasts and osteoclasts determines the mass and density of bone [1].
Studies of the mechanisms underlying osteogenesis have been greatly facilitated by the use of the undifferentiated preosteoblastic cell line MC3T3-E1, available from ATCC. This cell line was derived from mouse calvaria [3,4] and under defined culture conditions can be induced to undergo a developmental sequence leading to the formation of multilayered bone nodules [4,5]. This sequence is characterized by the replication of preosteoblasts followed by growth arrest and expression of mature osteoblastic characteristics such as matrix maturation and eventual formation of multilayered nodules with a mineralized extracellular matrix [5]. This cell line has become the standard in vitro model of osteogenesis and has found widespread use in studies examining many aspects and applications of osteogenesis, including transcriptional regulation [5], mineralization [3,6] and tissue engineering [7].
Millipore's In Vitro Osteogenesis Assay Kit provides the necessary reagents and protocols for efficient in vitro differentiation of MC3T3-E1 cells to mature osteoblasts, and for the end-point determination and quantification of this process.
Application:
Millipore's In Vitro Osteogenesis Assay Kit contains all reagents necessary to efficiently differentiate MC3T3-E1cells to a mature osteoblastic lineage, as determined by staining for mineralization. In addition this kit provides all the necessary reagents and a protocol for quantifying osteogenesis using a standard plate reader. This product is useful for studying the effects of growth factors, drugs and toxic agents on bone formation and for screening candidate osteogenic pharmaceuticals. Also, studies examining the intracellular signaling pathways regulating osteoblast differentiation can be facilitated by use of this kit.
The kit includes the osteogenesis-inducing reagents: ascorbic acid 2-phosphate, β-glycerophosphate and melatonin, and Alizarin Red Solution, a staining solution used to detect the presence of calcium in bone. Additionally, we provide reagents and a protocol for performing quantitative analysis of Alizarin Red staining.
Using Millipore's In Vitro Osteogenesis Assay Kit, we routinely obtain >75% mature osteoblasts from a starting population of uninduced MC3T3-E1 cells obtained from ATCC. Efficiency of osteoblastic differentiation may vary, depending upon the quality of the cells used, and if variatio |