Green fluorescent protein (GFP) is a protein, comprised of 238 amino acids (26,9 kDa), originally isolated from the jellyfish Aequorea victoria that fluoresces green when exposed to blue light. The GFP gene can be introduced into organisms and maintained in their genome through breeding, or local injection. Find GFP assays here to detect the amount of GFP present in your experiment.
Constructing and Expressing GFP Fusion Proteins The following protocol can be used for the development of stable cell lines expressing GFP fusion proteins. Although optimal transfection procedures (e.g., calcium phosphate, electroporation, or FuGENE 6 [Roche Applied Science]) vary depending on cell type, this general transfection procedure has been successful for stable transfection of HeLa, A-431, U2OS, BHK, and HT1080 cells.
Imaging Hoechst-Labeled Chromosomes and Fluorescent Proteins during the Cell Cycle Specific molecular components can be efficiently labeled by a combination of three methods: chemical transfection of GFP-fusion constructs, staining of chromosomes with the DNA-specific, fluorescent dye Hoechst 33342, and microinjection of fluorescently conjugated proteins. This procedure provides an example of using all three methods in sequence to label components of living HeLa cells. These methods should be followed in the order presented, but any of them can be omitted when not needed.
Monster Green® Fluorescent Protein Assay Green fluorescent protein is commonly used to monitor gene expression and protein trafficking within intact cells. The Monster Green® Fluorescent Protein is encoded by an improved synthetic version of the green fluorescent protein gene originally cloned from Montastrea cavernosa (Great Star Coral).
Probing Protein Interactions Using GFP and FRET Protocol specifically describes data acquisition for a particular variant of GFP (EGFP) or Oregon Green as a donor fluorophore, but it can be adapted for image acquisition of other chromophore.
Probing Protein Interactions Using GFP and FRET Protocol In preparation for FLIM-FRET analysis, the appropriate donor and acceptor components must be introduced into live or fixed cells. The method of introduction depends on the nature of the components and the state of the cells. For example, plasmid DNAs encoding a protein of interest fused to a variant of GFP may be introduced into live cells by transfection or microinjection, whereas labeled antibodies are delivered by microinjection.