Journal
BIOTECHNIQUES
Volume 44, Issue 1, Pages 70-+Publisher
BIOTECHNIQUES OFFICE
DOI: 10.2144/000112685
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We have developed a set of simple modifications of the green fluorescent protein (GFP)fragment reassembly assay in bacteria that permits: (i) fluorescent microscopy visualization of GFP reassembly only 1-2 h after induction of protein expression, thus approximating the detection of GFP reassembly to the real-time dynamics of protein complex formation in living cells; (ii) spectrofluorometric detection of reassembled GFP fluorescent signals directly in lysates from cell suspension thereby avoiding, in many cases, the need for tag-affinity isolation of protein complexes; and (iii) comparative quantification of signal intensity in numerous cell-sample lysates using a Bio-Rad IQ5 spectrofluorometric detection system (Bio-Rad Laboratories, Madrid, Spain). Collectively, the results demonstrate that the combination of microscopic and spectrofluorometric detection provides a time-saving and sensitive alternative to existing methods of fluorescence complementation analysis.
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