hCMEC/D3 Cells

Categories: Transformed cell lines Description: The HCMEC/D3 cell line represents an immortalized human cerebral microvascular endothelial cell line, extensively utilized in the study of the blood-brain barrier (BBB). This cell line was generated through the transduction of primary human cerebral microvascular endothelial cells with a lentiviral vector expressing human telomerase reverse transcriptase (hTERT), a crucial enzyme for maintaining telomere length and thereby promoting cellular longevity without transforming the cell phenotype. The introduction of hTERT helps these cells to bypass the replicative senescence that limits the lifespan of primary cells, allowing sustained propagation in culture. HCMEC/D3 cells retain key physiological and morphological characteristics of primary cerebral endothelial cells, making them a valuable model for in vitro studies of the BBB. These include the expression of tight junction proteins such as claudin-5, occludin, and zonula occludens-1, which are critical for maintaining barrier integrity. The cells also express various transporters and receptors typical of the cerebral endothelium, supporting their use in studies related to drug delivery and neurovascular disorders. The ability of HCMEC/D3 to form a tight monolayer with high electrical resistance underscores their suitability for BBB permeability assays. Research utilizing HCMEC/D3 cells has covered a wide range of applications, including the investigation of cerebral pathologies such as stroke, multiple sclerosis, and metastasis of cancer to the brain. Their compatibility with various molecular biology techniques also makes them an excellent tool for studying endothelial cell responses to inflammatory stimuli, shear stress, and neurotoxic substances. This cell line provides a robust, reproducible platform for dissecting the molecular events at the cerebral endothelial level, contributing valuable insights into the complexities of neurovascular health and disease. Organism: Human Tissue: Brain, temporal lobe, blood microvessel Synonyms: HCMEC/D3, CMEC/D3, human Cortical Microvessels Endothelial Cells/D3 Age: Adult Gender: Female Morphology: Endothelial Cell Type: Endothelial cell Growth Properties: Adherent Citation: hCMEC/D3 (Cytion catalog number 305024) Biosafety Level: 1 Ncbi_ Taxid: 9606.0 Cellosaurus Accession: CVCL_U985 Viruses: Transformant: Simian virus 40 (SV40) Culture Medium: EGM -2 MV Microvascular Endothelial Cell Growth Medium-2 BulletKit (from Lonza, Lonza catalog number CC-3202) Supplements: Supplement the supplied EBM-2 Basal Medium as recommended by the manufacturer Freeze Medium: As a cryopreservation medium, use 50% basal medium + 40% FBS + 10% DMSO, or CM-1 (Cytion catalog number 800100), which includes optimized osmoprotectants and metabolic stabilizers to enhance recovery and reduce cryo-induced stress. Thawing And Culturing Cells: Confirm that the vial remains deeply frozen upon delivery, as cells are shipped on dry ice to maintain optimal temperatures during transit. Upon receipt, either store the cryovial immediately at temperatures below -150°C to ensure the preservation of cellular integrity, or proceed to step 3 if immediate culturing is required. For immediate culturing, swiftly thaw the vial by immersing it in a 37°C water bath with clean water and an antimicrobial agent, agitating gently for 40-60 seconds until a small ice clump remains. Perform all subsequent steps under sterile conditions in a flow hood, disinfecting the cryovial with 70% ethanol before opening. Carefully open the disinfected vial and transfer the cell suspension into a 15 ml centrifuge tube containing 8 ml of room-temperature culture medium, mixing gently. Centrifuge the mixture at 300 x g for 3 minutes to separate the cells and carefully discard the supernatant containing residual freezing medium. Gently resuspend the cell pellet in 10 ml of fresh culture medium. For adherent cells, divide the suspension between two T25 culture flasks, for suspension cultures, transfer all the medium into one T25 flask to promote effective cell interaction and growth. Adhere to established subculture protocols for continued growth and maintenance of the cell line, ensuring reliable experimental outcomes. Sterility: Mycoplasma contamination is excluded using both PCR-based assays and luminescence-based mycoplasma detection methods. To ensure there is no bacterial, fungal, or yeast contamination, cell cultures are subjected to daily visual inspections. Safety Precautions: When planning to store a cryovial in liquid nitrogen for future thawing, it is mandatory to adhere to stringent safety measures. Appropriate protective gloves and clothing are essential, and the use of a face mask or safety goggles is required during the transfer of frozen samples to or from the liquid nitrogen tank. This is to mitigate the risk of injury from potential cryovial explosions upon removal, which can result in the projection of sharp fragments. Warranty: We stand by the promise of delivering products with high cell viability and robust culture performance. To achieve the best results, please make sure you follow the storage and culture instructions detailed in the product information sheet closely. Your adherence to these guidelines is key to success. Subject To Material Transfer Agreements: If you intend to use Cytion cell lines solely for internal research at a single research site, please complete and sign our Material Transfer Agreement (MTA) and submit it along with your order.For any commercial applications - including but not limited to fee-for-service work, quality control testing, product release, diagnostic use, or regulatory studies - please complete the Intended Use Form so we can prepare a suitable agreement tailored to your project.Please note: The MTA applies only to certain cell lines. If this notice and the MTA document appear on a product page, the agreement is applicable. For cell lines not covered by the MTA, no reference to the agreement will be shown. The MTA is not valid for customers in the Americas, China, or Taiwan. Please contact our U.S. entity to receive the appropriate agreement. Required Product 3: 860015.0 Required Product 4: 830100.0