LentiBrite GfpControlLentiviralBiosensor

Stock Code: 3586380
Manufacturer Part No: 17-10387

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Descriptions

Application

Research Sub CategoryAll

Research CategoryCell Structure

Fluorescence Microscopy
Imaging:
(See Figure 1 in datasheet)
HeLa cells were plated in a chamber slide and transduced with lentiviral particles at an MOI of 40 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 displays a uniform green fluorescent distribution in the cells.

Immunocytochemistry Comparison:
(See Figure 2 in datasheet)
Similar to Figure 1 (see datasheet), U2OS cells were plated in a chamber slide and transduced with lentiviral particles at an MOI of 40 for 24 hours. After 24 hours, media was replaced and cells were then further incubated for 48 hours.

Hard-to-transfect Cell Types:
(See Figure 3 in datasheet)
Primary cell types HUVEC or HuMSC were plated in chamber slides and transduced with lentiviral particles at an MOI of 40 for 24 hours.

Additional cell type:
(See Figure 4 in datasheet)
Primary Rat Cortical Neuron cells were treated as in Figure 1 (see datasheet).

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.

Components

TagGFP2 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, human mesenchymal stem cells (HuMSC)), while others may require higher MOIs (e.g., human umbilical vein endothelial cells (HUVEC)).
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.

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 HereBiosensors 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

Specifications

Quality Level	100
Manufacturer	SIGMA-ALDRICH
Technique(s)	immunocytochemistry: suitable, cell based assay: suitable, transfection: suitable, immunofluorescence: suitable