Fluorescent labeling of antibody fragments using split GFP.
Fluorescent labeling of antibody fragments using split GFP.
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DOI：
10.1371/journal.pone.0025727
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发表时间：
2011
期刊：
影响因子：
3.7
通讯作者：
Bradbury AR
中科院分区：
文献类型：
--
作者：
Ferrara F;Listwan P;Waldo GS;Bradbury AR
Antibody fragments are easily isolated from in vitro selection systems, such as phage and yeast display. Lacking the Fc portion of the antibody, they are usually labeled using small peptide tags recognized by antibodies. In this paper we present an efficient method to fluorescently label single chain Fvs (scFvs) using the split green fluorescent protein (GFP) system. A 13 amino acid tag, derived from the last beta strand of GFP (termed GFP11), is fused to the C terminus of the scFv. This tag has been engineered to be non-perturbing, and we were able to show that it exerted no effect on scFv expression or functionality when compared to a scFv without the GFP11 tag. Effective functional fluorescent labeling is demonstrated in a number of different assays, including fluorescence linked immunosorbant assays, flow cytometry and yeast display. Furthermore, we were able to show that this split GFP system can be used to determine the concentration of scFv in crude samples, as well an estimate of antibody affinity, without the need for antibody purification. We anticipate this system will be of widespread interest in antibody engineering and in vitro display systems.
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DOI：
10.1007/s10969-005-5247-5
发表时间：
2005-09-01
期刊：
Journal of Structural and Functional Genomics
影响因子：
--
作者：
Cabantous, Stephanie;Pedelacq, Jean-Denis;Waldo, Geoffrey S.
通讯作者：
Waldo, Geoffrey S.
影响因子：
2.2
作者：
Morino, K;Katsumi, H;Kurosawa, Y
通讯作者：
Kurosawa, Y
影响因子：
46.9
作者：
Cabantous, S;Terwilliger, TC;Waldo, GS
通讯作者：
Waldo, GS
影响因子：
1.6
作者：
Griep, RA;van Twisk, C;Schots, A
通讯作者：
Schots, A
影响因子：
4.8
作者：
Hink, MA;Griep, RA;Visser, AJWG
通讯作者：
Visser, AJWG