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17-10147 LentiBrite™ GFP-HMGB1 Lentiviral Biosensor

LentiBrite™ GFP-HMGB1 Lentiviral Biosensor MSDS (material safety data sheet) or SDS, CoA and CoQ, dossiers, brochures and other available documents.
Key Applications: Transfection, Immunofluorescence, Immunocytochemistry
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17-10147
1 vial (minimum of 3 x 10E8 IFU/mL)  
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      Overview

      Replacement Information

      Key Specifications Table

      Key ApplicationsDetection Methods
      TFX, IF, ICCFluorescent
      Description
      Catalogue Number17-10147
      Trade Name
      • LentiBrite
      • Chemicon
      DescriptionLentiBrite™ GFP-HMGB1 Lentiviral Biosensor
      OverviewRead our application note in Nature Methods!
      http://www.nature.com/app_notes/nmeth/2012/121007/pdf/an8620.pdf
      (Click Here!)

      Learn more about the advantages of our LentiBrite Lentiviral Biosensors! Click Here

      Biosensors can be used to detect the presence/absence of a particular protein as well as the subcellular location of that protein within the live state of a cell. Fluorescent tags are often desired as a means to visualize the protein of interest within a cell by either fluorescent microscopy or time-lapse video capture. Visualizing live cells without disruption allows researchers to observe cellular conditions in real time.

      Lentiviral vector systems are a popular research tool used to introduce gene products into cells. Lentiviral transfection has advantages over non-viral methods such as chemical-based transfection including higher-efficiency transfection of dividing and non-dividing cells, long-term stable expression of the transgene, and low immunogenicity.

      EMD Millipore is introducing LentiBrite™ Lentiviral Biosensors, a new suite of pre-packaged lentiviral particles encoding important and foundational proteins of autophagy, apoptosis, and cell structure for visualization under different cell/disease states in live cell and in vitro analysis.
      • Pre-packaged, fluorescently-tagged with GFP & RFP
      • Higher efficiency transfection as compared to traditional chemical-based and other non-viral-based transfection methods
      • Ability to transfect dividing, non-dividing, and difficult-to-transfect cell types, such as primary cells or stem cells
      • Non-disruptive towards cellular function

      EMD Millipore’s LentiBrite™ GFP-HMGB1 lentiviral particles provide bright fluorescence and precise localization to enable live cell analysis of HMGB1 translocation in difficult-to-transfect cell types.
      Alternate Names
      • High mobility group protein B1
      • High mobility group protein 1
      Background InformationHMBG1 (high mobility box group 1) functions in the nucleus of most cells to stabilize nucleosomes and enhance binding of transcription factor complexes to DNA. In certain activated inflammatory cells, HMGB1 is actively secreted, whereas necrotic cells passively release HMGB1. In both cases, extracellular HMGB1 can bind to multiple immune cell types via RAGE (receptor for advanced glycation end products) to stimulate chemotaxis and cytokine production, which, depending on the context, results in inflammation, wound healing, or immunogenic tumor cell death. Cells undergoing autophagy also display translocation of HMGB1, from the nucleus to the cytoplasm. Expression of genetic fusions between HMGB1 and fluorescent proteins permits monitoring of HMGB1 translocation in real time.
      EMD Millipore’s LentiBrite™ GFP-HMGB1 lentiviral particles provide bright fluorescence and precise localization to enable live cell analysis of HMGB1 translocation in difficult-to-transfect cell types.
      References
      Product Information
      Components
      • TagGFP2-HMGB Lentivirus:
        One vial containing 25 µL of lentiviral particles at a minimum of 3 x 10E8 infectious units (IFU) per mL.
        For lot-specific titer information, please see lot specific “Viral Titer” in the product specifications of the datasheet.

      • Promoter
        EF-1 (Elongation Factor-1)

      • Multiplicty of Infection (MOI)
        MOI = Ratio of # of infectious lentiviral particles (IFU) to # of cells being infected.
        Typical MOI values for high transduction efficiency and signal intensity are in the range of 20-40. For this target, some cell types may require lower MOIs (e.g., HT-1080, HeLa, U2OS), while others may require higher MOIs (e.g., human umbilical vein endothelial cells (HUVEC), human mesenchymal stem cells (HuMSC)).
        NOTE: MOI should be titrated and optimized by the end user for each cell type and lentiviral target to achieve desired transduction efficiency and signal intensity.
      Detection methodFluorescent
      Quality LevelMQ100
      Applications
      Key Applications
      • Transfection
      • Immunofluorescence
      • Immunocytochemistry
      Application NotesFluorescence Microscopy Imaging:
      (See Figure 1 in datasheet)
      U2OS cells were plated in a chamber slide and transduced with lentiviral particles at an MOI of 10 for 24 hours. After media replacement and 48 hours further incubation, cells were fixed with formaldehyde and mounted. Images were obtained by oil immersion wide-field fluorescence microscopy. The GFP-HMGB1 displays in the nucleus of cells.

      Immunocytochemistry Comparison and Inhibitor Analysis:
      (See Figure 2 in datasheet)
      Similar to Figure 1 (See datasheet), HeLa cells were plated in a chamber slide and transduced with lentiviral particles at an MOI of 10 for 24 hours. After 24 hours, media was replaced and cells were then further incubated for 48 hours. Immunocytochemical staining (blue) of the same fields of view with a nuclear stain (DAPI) reveals similar expression patterns to the protein-GFP (green).

      Difficult-to-transfect Cell Types:
      (See Figure 3 in datasheet)
      Primary cell types HUVEC and Human mesenchymal stem cells (HuMSC) were plated in chamber slides and transduced with lentiviral particles at an MOI of 20 for 24 hours.

      For optimal fluorescent visualization, it is recommended to analyze the target expression level within 24-48 hrs after transfection/infection for optimal live cell analysis, as fluorescent intensity may dim over time, especially in difficult-to-transfect cell lines. Infected cells may be frozen down after successful transfection/infection and thawed in culture to retain positive fluorescent expression beyond 24-48 hrs. Length and intensity of fluorescent expression varies between cell lines. Higher MOIs may be required for difficult-to-transfect cell lines.
      Biological Information
      Gene Symbol
      • HMGB1
      • HMG1
      • HMGB
      • HMG-1
      Purification MethodPEG precipitation
      UniProt Number
      Physicochemical Information
      Dimensions
      Materials Information
      Toxicological Information
      Safety Information according to GHS
      Safety Information
      Product Usage Statements
      Quality AssuranceEvaluated by transduction of HT-1080 cells and fluorescent imaging performed for assessment of target localization and transduction efficiency.
      Usage Statement
      • This product contains genetically modified organisms (GMO). Within the EU GMOs are regulated by Directives 2001/18/EC and 2009/41/EC of the European Parliament and of the Council and their national implementation in the member States respectively. This legislation obliges MilliporeSigma to request certain information about you and the establishment where the GMOs are being handled. Click here for Enduser Declaration (EUD) Form.
      • Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.
      Storage and Shipping Information
      Storage ConditionsStorage and Handling
      Lentivirus is stable for at least 4 months from date of receipt when stored at -80°C. After first thaw, place immediately on ice and freeze in working aliquots at -80°C. Frozen aliquots may be stored for at least 2 months. Further freeze/thaws may result in decreased virus titer and transduction efficiency.

      IMPORTANT SAFETY NOTE
      Replication-defective lentiviral vectors, such as the 3rd Generation vector provided in this product, are not known to cause any diseases in humans or animals. However, lentiviruses can integrate into the host cell genome and thus pose some risk of insertional mutagenesis. Material is a Risk Group 2 and should be handled under BSL2 controls. A detailed discussion of biosafety of lentiviral vectors is provided in Pauwels, K. et al. (2009). State-of-the-art lentiviral vectors for research use: Risk assessment and biosafety recommendations. Curr. Gene Ther. 9: 459-474.
      Packaging Information
      Material Size1 vial (minimum of 3 x 10E8 IFU/mL)
      Transport Information
      Supplemental Information
      Specifications
      Global Trade Item Number
      Catalog Number GTIN
      17-10147 04053252549205

      Documentation

      LentiBrite™ GFP-HMGB1 Lentiviral Biosensor SDS

      Title

      Safety Data Sheet (SDS) 

      LentiBrite™ GFP-HMGB1 Lentiviral Biosensor Certificates of Analysis

      TitleLot Number
      LentiBrite GFP-HMGB1 Lentiviral Biosensor - 2201533 2201533
      LentiBrite GFP-HMGB1 Lentiviral Biosensor - Q1988426 Q1988426
      LentiBrite™ GFP-HMGB1 -2567162 2567162
      LentiBrite™ GFP-HMGB1 Lentiviral Biosensor - 3212812 3212812
      LentiBrite™ GFP-HMGB1 Lentiviral Biosensor - 3429398 3429398
      LentiBrite™ GFP-HMGB1 Lentiviral Biosensor - 3599015 3599015
      LentiBrite™ GFP-HMGB1 Lentiviral Biosensor - 3698224 3698224
      LentiBrite™ GFP-HMGB1 Lentiviral Biosensor - 3979360 3979360
      LentiBrite™ GFP-HMGB1 Lentiviral Biosensor - 4034280 4034280
      LentiBrite™ GFP-HMGB1 Lentiviral Biosensor Packaged Lentiviral Particles 3020581