Characterization of a chimeric aequorin molecule expressed in myeloma cells.
We have constructed a chimeric aequorin consisting of a fragment of the anti-NP immunoglobulin gene fused to the aequorin gene. Expression in a myeloma cell line has produced a Fab'-like molecule that has the ability to bind NIP specifically and generate bioluminescent activity. It takes approximately 8 h at 4 degrees C in the presence of 2-mercaptoethanol and coelenterazine to regenerate luminescent activity. While the flash kinetics of this recombinant molecule are similar to native aequorin, its quantum efficiency is ten times lower. Preliminary studies have been conducted to ascertain its usefulness for immunoassays. We have shown for this chimeric aequorin 7 x 10(-19) moles can be detected in solution, also it can be used in a solid-phase assay and is stably stored at -70 degrees C for at least 2 months.
Casadei
J
J
IGEN Inc., Rockville, MD 20852.
Powell
M J
MJ
Kenten
J H
JH
D000331:Aequorin / Q000235:genetics* / Q000378:metabolism; D000818:Animals; D003001:Cloning, Molecular; D004355:Drug Stability; D005786:Gene Expression Regulation; D007700:Kinetics; D008163:Luminescent Measurements; D008164:Luminescent Proteins / Q000235:genetics*; D009101:Multiple Myeloma / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014407:Tumor Cells, Cultured / Q000378:metabolism
D008164:Luminescent Proteins; D011994:Recombinant Proteins; D000331:Aequorin