Development of an open sandwich fluoroimmunoassay based on fluorescence resonance energy transfer.
We have developed a sensitive, one-step, homogeneous open sandwich fluoroimmunoassay (OsFIA) based on fluorescence resonance energy transfer (FRET) and luminescent semiconductor quantum dots (QDs). In this FRET assay, estrogen receptor beta (ER-beta) antigen was incubated with QD-labeled anti-ER-beta monoclonal antibody and Alexa Fluor (AF)-labeled anti-ER polyclonal antibody for 30 min, followed by FRET measurement. The dye separation distance was estimated between 80 and 90 A. The current method is rapid, simple, and highly sensitive, and it did not require the bound/free reagent separation steps and solid-phase carriers. A concentration as low as 0.05 nM (2.65 ng/ml) receptor was detected with linearity. In addition, the assay was performed with commercial antibodies. This assay provides a convenient alternative to conventional, laborious sandwich immunoassays.
Wei
Quande
Q
Department of Pathogenic Biology, Sun Yat-sen University, Guangzhou 530080, People's Republic of China.
Lee
Moonkwon
M
Yu
Xinbing
X
Lee
Eun Kyu
EK
Seong
Gi Hun
GH
Choo
Jaebum
J
Cho
Yong Woo
YW
D000911:Antibodies, Monoclonal; D047629:Estrogen Receptor beta / Q000032:analysis* / Q000276:immunology; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D005456:Fluorescent Dyes; D015200:Fluoroimmunoassay / Q000379:methods*; D008956:Models, Chemical; D045663:Quantum Dots; D012680:Sensitivity and Specificity
D000911:Antibodies, Monoclonal; D047629:Estrogen Receptor beta; D005456:Fluorescent Dyes