The green fluorescent protein (GFP) and its mutants have been extensively used to study various cellular processes and, more recently, as labels in binding assays. We have employed a mutant of GFP, an enhanced GFP (EGFP), in the development of homogeneous assays for biotin and cortisol. To demonstrate the feasibility of using EGFP as a label with different kinds of binders in the development of homogeneous assays, we employed the biotin-avidin and an antigen-antibody as the binding pairs. Biotin and cortisol were chemically conjugated to EGFP. A quenching of fluorescence intensity of EGFP was observed upon binding of avidin to the EGFP-biotin conjugate. The percentage fluorescence quenching observed decreased as the concentration of free biotin in the sample increased due to the fewer binding sites on avidin available for binding to the EGFP-biotin conjugate. A dose-response curve for biotin was generated by relating percentage fluorescence quenched with free biotin in the sample. To determine whether EGFP can undergo a similar type of homogeneous change when used as a label for antigen-antibody type of binding, cortisol was selected as a model analyte. In the presence of an anti-cortisol antibody the fluorescence signal of the EGFP-cortisol conjugate was quenched. A dose-response curve for cortisol was generated by relating the quenching in the fluorescence signal with varying amounts of free cortisol in the sample. This is the first time that GFP or one of its mutants has been employed as a label in homogeneous assays, thus enhancing the versatility of employing GFP or its mutants in a number of bioanalytical applications, such as clinical analysis and high-throughput screening systems.
This research output has an Altmetric Attention Score of 6. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 18 January 2011.
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#7,405,506
of 34,043,423 outputs
Outputs from Analytical Biochemistry
#2,331
of 10,879 outputs
Outputs of similar age
#21,816
of 160,324 outputs
Outputs of similar age from Analytical Biochemistry
#22
of 57 outputs
Altmetric has tracked 34,043,423 research outputs across all sources so far. This one has received more attention than most of these and is in the 73rd percentile.
So far Altmetric has tracked 10,879 research outputs from this source. They typically receive a little more attention than average, with a mean Attention Score of 5.2. This one has gotten more attention than average, scoring higher than 58% of its peers.
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We're also able to compare this research output to 57 others from the same source and published within six weeks on either side of this one. This one is in the 43rd percentile – i.e., 43% of its contemporaries scored the same or lower than it.
