A radioisotope label-free alpha-bungarotoxin-binding assay using BIAcore sensor chip technology for real-time analysis.
alpha-Bungarotoxin (alpha-bgtx)-binding proteins, including certain nicotinic acetylcholine receptors and acetylcholine-binding proteins (AChBPs), are frequently characterized with radioisotope-labeled alpha-bgtx-binding assays. Such assays, however, preclude investigations of binding interactions in real time and are hampered by the inconveniences associated with radioisotope-labeled reagents. We used surface plasmon resonance-based technology (BIAcore) to investigate the binding of recombinant AChBP to CM-5 sensor chip surfaces with directly immobilized alpha-bgtx. We validated our BIAcore results by comparing the same biological samples using the traditional (125)I-labeled alpha-bgtx-binding assay. An alpha-bgtx sensor chip, as described here, enables detailed, real-time, radioisotope-free interaction studies that can greatly facilitate the characterization of novel alpha-bgtx-binding proteins and complexes.
Paulo
Joao A
JA
Graduate Program in Molecular Biology, Cell Biology, and Biochemistry, Brown University, Providence, RI 02912, USA.
Hawrot
Edward
E
D015374:Biosensing Techniques / Q000379:methods*; D002038:Bungarotoxins / Q000032:analysis* / Q000737:chemistry*; D002352:Carrier Proteins / Q000032:analysis; D007553:Isotope Labeling; D011868:Radioisotopes; D011978:Receptors, Nicotinic / Q000032:analysis; D020349:Surface Plasmon Resonance / Q000379:methods*
D002038:Bungarotoxins; D002352:Carrier Proteins; D011868:Radioisotopes; D011978:Receptors, Nicotinic