Phycobiliproteins are fluorescent proteins derived from cyanobacteria and eukaryotic algae. Their fluorescence is much higher than chemical fluorescent probes such as fluorescein and rhodamine. R-phycoerythrin (R-PE) is one of the phycobiliproteins and has an orange fluorescence at around 578 nm, and it can be excited at 488 nm (Fig. 1). Because of this high fluorescence, phycobiliprotein-labeled antibodies or other molecules can give greater sensitivity in flow cytometry and immunostaining. R-phycoerythrin Labeling Kit-SH is for simple and rapid preparation of R-PE-labeled IgG (Fig. 2). SH-reactive R-PE (a component of this kit) has a maleimide group and can easily make a covalent bond with a sulfhydryl group of the target molecule without any activation process. The filtration tube in this kit allows a quick buffer exchange and concentration of sample IgG solution. This kit contains all the reagents necessary for R-PE labeling, including the reducing agent for preparation of reduced IgG that has an SH group and the storage buffer for conjugates.
Fig. 1 Fluorescence spectrum of R-PE
Excitation wavelength: 566 nm
Emission wavelength: 578 nm
Fig. 2 IgG labeling reaction of SH-reactive R-PE
♦ The molecular weight of the reduced protein to be labeled with this kit should be greater than 50,000.
♦ IgG or R-Phycoerythrin-conjugated IgG is always on the membrane of the filtration tube during the labeling process.
♦ If the IgG solution contains other proteins with molecular weight larger than 10,000, such as BSA or gelatin, purify the IgG solution prior to labeling R-phycoerythrin with this kit. IgG solution can be purified by IgG Purification Kits (not included in this kit).
♦ If the IgG solution contains small insoluble materials, centrifuge the solution and use the supernatant for the labeling.
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