Direct, ultrasensitive, and selective optical detection of protein toxins using multivalent interactions.
Three highly sensitive, selective, and reagent-free optical signal transduction methods for detection of polyvalent proteins have been developed by directly coupling distance-dependent fluorescence self-quenching and/or resonant-energy transfer to the protein-receptor binding events. The ganglioside GM1, as the recognition unit for cholera toxin (CT), was covalently labeled with fluorophores and then incorporated into a biomimetic membrane surface. The presence of CT with five binding sites for GM1 causes dramatic change for the fluorescence of the labeled GM1. (1) In the scheme using fluorescence self-quenching as a signal-transduction mechanism, the fluorescence intensity drops significantly as a result of aggregation of the fluorophore-labeled GM1 on a biomimetic surface. (2) By labeling GM1 with a fluorescence energy transfer pair, aggregation of the labeled GM1 results in a decrease in donor fluorescence and an increase in acceptor fluorescence, providing a unique signature for selective protein-receptor binding. (3) In the third scheme, using the biomimetic surface as part of signal transduction and combining both fluorescence self-quenching and energy-transfer mechanisms to enhance the signal transduction, a signal amplification was achieved. The detection systems can reliably detect less than 0.05 nM CT with fast response (less than 5 min). This approach can easily be adapted to any biosensor scheme that relies on multiple receptors or co-receptors. The methods can also be applied to investigate the kinetics and thermodynamics of the multivalent interactions.
Song
X
X
Chemical Science and Technology Division, Los Alamos National Laboratory, New Mexico 87545, USA. xs.xsong@lanl.gov
Swanson
B I
BI
D000418:Albumins / Q000737:chemistry; D001665:Binding Sites; D001896:Boron Compounds / Q000737:chemistry; D002772:Cholera Toxin / Q000032:analysis / Q000737:chemistry / Q000378:metabolism; D003287:Contrast Media / Q000737:chemistry; D004735:Energy Transfer; D019793:Fluorescein / Q000032:analysis / Q000737:chemistry; D005456:Fluorescent Dyes / Q000737:chemistry / Q000378:metabolism; D005677:G(M1) Ganglioside / Q000737:chemistry / Q000378:metabolism; D008051:Lipid Bilayers; D008567:Membranes, Artificial; D055095:Optics and Photonics; D010713:Phosphatidylcholines / Q000737:chemistry; D011506:Proteins / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism*; D012680:Sensitivity and Specificity; D015398:Signal Transduction; D013050:Spectrometry, Fluorescence / Q000295:instrumentation / Q000379:methods*; D014118:Toxins, Biological / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism
C095489:4,4-difluoro-4-bora-3a,4a-diaza-s-indacene; D000418:Albumins; D001896:Boron Compounds; D003287:Contrast Media; D005456:Fluorescent Dyes; D008051:Lipid Bilayers; D008567:Membranes, Artificial; D010713:Phosphatidylcholines; D011506:Proteins; D014118:Toxins, Biological; D005677:G(M1) Ganglioside; D002772:Cholera Toxin; C028694:1-palmitoyl-2-oleoylphosphatidylcholine; D019793:Fluorescein