Saltar al contenido
Merck
Todas las fotos(5)

Key Documents

17-10153

Sigma-Aldrich

LentiBrite RFP-Vimentin Lentiviral Biosensor

Iniciar sesiónpara Ver la Fijación de precios por contrato y de la organización


About This Item

UNSPSC Code:
12352207
eCl@ss:
34360190
NACRES:
NA.51

manufacturer/tradename

Chemicon®
LentiBrite

Quality Level

technique(s)

cell based assay: suitable
immunocytochemistry: suitable
immunofluorescence: suitable
single cell analysis: suitable
transfection: suitable

UniProt accession no.

detection method

fluorometric

shipped in

dry ice

Gene Information

human ... VIM(7431)

General description

Read 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 RFP-Vimentin lentiviral particles provide bright fluorescence and precise localization to enable live cell analysis of vimentin IF dynamics in difficult-to-transfect cell types.
Intermediate filaments (IFs) are 10 nm diameter cytoskeletal structures that typically form a cage-like structure extending from the nucleus to the plasma membrane. The five families of IF proteins have distinct polymer characteristics and cell type-specific expression patterns. Expression of vimentin, a type-III IF protein, is restricted to mesenchymal cells, and appearance of vimentin in epithelial cancer cells is a hallmark of the epithelial-to-mesenchymal transition. Vimentin assembles into classic long filaments and short filaments termed squiggles. Time-lapse microscopy of live cells expressing RFP-vimentin has revealed the dynamics of vimentin IF maturation, and crosstalk with microtubules and microfilaments.
EMD Millipore’s LentiBrite Vimentin-RFP lentiviral particles provide bright fluorescence and precise localization to enable live cell analysis of vimentin IF dynamics in difficult-to-transfect cell types.

Application

Immunocytochemistry Comparison Analysis:
Similar to Figure 1 in the datasheet, HeLa cells were plated in a chamber slide and transduced with lentiviral particles at an MOI of 20 for 24 hours. After 24 hours, media was replaced and cells were then further incubated for 48 hours. Immunocytochemical staining (green) of the same fields of view with a monoclonal antibody against Vimentin (Cat No. MAB3400) reveals similar expression patterns to the RFP-protein (red).

Hard-to-transfect Cell Types:
Primary cell types HUVEC or HuMSC were plated in chamber slides and transduced with lentiviral particles at an MOI of 20 and 10, respectively, for 24 hours.

Confocal Microscopy Imaging: HeLa and U2OS cells were treated as in Figure 2A in the datasheet. Images were obtained by oil immersion confocal fluorescence microscopy.

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.
Research Category
Cell Structure
Research Sub Category
Cytoskeleton

Components

TagRFP-Vimentin 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 “Viral Titer” in the product specifications above.


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), while others may require higher MOIs (e.g., human mesenchymal stem cells (HuMSC), human umbilical vein endothelial cells (HUVEC ), HeLa, U2OS).
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.

Quality

Evaluated by transduction of HT-1080 cells and fluorescent imaging performed for assessment of transduction efficiency.

Physical form

PEG precipitation

Storage and Stability

Storage 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.

Legal Information

CHEMICON is a registered trademark of Merck KGaA, Darmstadt, Germany

Storage Class

10 - Combustible liquids

wgk_germany

WGK 2


Certificados de análisis (COA)

Busque Certificados de análisis (COA) introduciendo el número de lote del producto. Los números de lote se encuentran en la etiqueta del producto después de las palabras «Lot» o «Batch»

¿Ya tiene este producto?

Encuentre la documentación para los productos que ha comprado recientemente en la Biblioteca de documentos.

Visite la Librería de documentos

Contenido relacionado

Fluorescent lentiviral particles encoding important GFP/RFP fusion proteins related to autophagy, apoptosis, and cell structure that enables live cell imaging.

Fluorescent lentiviral particles encoding important GFP/RFP fusion proteins related to autophagy, apoptosis, and cell structure that enables live cell imaging.

Fluorescent lentiviral particles encoding important GFP/RFP fusion proteins related to autophagy, apoptosis, and cell structure that enables live cell imaging.

Fluorescent lentiviral particles encoding important GFP/RFP fusion proteins related to autophagy, apoptosis, and cell structure that enables live cell imaging.

Nuestro equipo de científicos tiene experiencia en todas las áreas de investigación: Ciencias de la vida, Ciencia de los materiales, Síntesis química, Cromatografía, Analítica y muchas otras.

Póngase en contacto con el Servicio técnico