Extragel Matrix instruction
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SCILIA Extragel Matrix Tel.: E-mail: Website: Product Introduction The basement membrane is a matrix under the basal surface of epithelial cells of animals. Scilia Extragel Matrix is a reconstituted matrix hydrogel formed by basement membrane components extracted from mouse tumor tissues. This matrix hydrogel is mainly composed of laminin, collagen IV, and heparan sulfate proteoglycans (Kleinman et al. 1986). Besides, it contains various growth factors, such as epidermal growth factor (EGF), platelet-derived growth factor (PDGF), nerve growth factor (NGF), basic fibroblast growth factor (FGF-2), transforming growth factor-β (TGF-β), and insulin-like growth factor (IGF) (Vu-kicevic et al. 1992). Product Characteristics Extragel Matrix is liquid at 4 ℃ but gel-like when heated to 37 ℃. This transformation phemomenon is reversible. It can be liquefied again when it is stored at 4℃ overnight. (Tip: It is recommended to store the Extragel Matrix in an ice box in a refrigerator at 4 ℃ to realize the full liquefaction of the reconstitute matrix hydrogel.) Storage Condition Extragel Matrix could be stored in a freezer at -80 ℃ for 2 years. Product Application Extragel Matrix can be applied to the growth, differentiation, metabolism and toxicology of organoids, in vivo and in vitro angiogenesis experiments, and tumor formation experiment of immunodeficient mice. Precautions Extragel Matrix would start solidifying after the temperature is higher than 10 ℃, so the operation should be performed on ice. The matrix hydrogel can be dissolved in basic culture medium precooled at 4 ℃, and the organoid can be released from the Extragel Matrix. Operation Method Organoid culture (1 hour) 1. Thaw the Extragel Matrix (Scilia-OM-1 or Scilia-OM-1-ph or Scilia-OM-3 or Scilia-OM-3-ph) in refrigerator at 4 ℃ overnight. 2. Preheat the 24-well plate in cell culture incubator. 3. Prepare aliquots of Extragel Matrix using precool tips. 4. Prepare the single cell pellet with 1×106 cells derived from patients or animal tissue, centrifuged at 300 g for 5 minutes. 5. Mix the cell pellet with 50 μL Extragel M

Add the mixture into the well of plate (50 μL per well). Keep the plate in incubator for 10 minutes, flip and keep after another 5 minutes. Add 500 μL culture medium to the well with matrix and cells. Change the medium every 3 days. Tumor Formation Experiment in Immunodeficient Mice (1 hour) 1. Mix 1×106 cells with Extragel Matrix (volume ratio=1:1) (Scilia-OM-2 or Scilia-OM-2-ph or Scilia-OM-5 or Scilia-OM-5-ph). The total volume of mixture for each injection should be 100 μL at least. 2. Inject the mixture of cells and Extragel Matrix with a 1 mL syringe mounted with a 16 g needle and...

temperature. 7. Remove diluted Extragel Matrix after coating and seed the hESC or ipsc cell solution. 8. Transfer the plate to incubator. Practical Application Cases Figure 1. Establishment of human bile duct organoids in the matrix hydrogel of Company A and Extragel Matrix, respectively. Human bile duct organoids are imaged after being stained with DAPI (nucleus, blue), anti-ZO-1 antibody (tight-junction protein), and Alexa Fluor 647 Phalloidin (cytoskeleton protein F-actin). Figure 2. Growth of human bile duct organoids derived from human embryonic stem cells in the matrix hydrogel of...

and Extragel Matrix for 13 days, respectively. Figure 4. Characterization of hepatocellular carcinoma organoids derived from patients in the matrix hydrogel of Company A and Extragel Matrix, with immunostaining by DAPI (nuclus), HNF4a (a hepatic biomarker) and GPC3 (hepatocellular carcinoma biomarker). Figure 5. Human umbilical vein endothelial cells (HUVECs) can form vascular networks in the matrix hydrogel of Company A and Extragel Matrix. HUVEC cells are imaged after being stained with living cell dye Calcein AM (green).

Figure 6. Rat neural stem cells can grow, differentiate and form neurite networks in the Extragel Matrix. Rat neural stem cells (NSCs) were cultured in Extragel Matrix for 7 days and imaged after being stained with neuron biomarker Tuj1 (red) and nuclear dye DAPI (blue) (A). The right figure presents the neuron with pseudo colour (B). This experiment can be used to evaluate neurotoxicity and the differentiation and development of neurons. Figure 7. Growth of hESCs on the matrix hydrogel of company A and OrganoGle for 2 days. Reference 1. Kleinman HK, et al, Basement membrane complexes with...