Dual-labeled time-resolved immunofluorometric assay for the determination of IgM antibodies to rubella virus and cytomegalovirus in human serum

Zhou, Jianwei; Lei, Lamei; Liang, Qian-Ni; Liu, Tiancai; Lin, Guanfeng; Dong, Zhi-Ning; Liang, Rong-Liang; Chen, Zhenhua; Wu, Yixuan

doi:10.1016/j.clinbiochem.2015.01.009

Cited by 8 publications

(2 citation statements)

References 30 publications

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“…15 Also, different lanthanide ions, such as Eu 3+ in combination with Sm 3+ , were used for multiplexed immunoassays. 16 However, all of these technologies are heterogeneous, which means that several, often timeconsuming, immobilization, washing, incubation, and separation steps are necessary for the complete assay procedure.…”

Section: ■ Introductionmentioning

confidence: 99%

“…Nanoparticles, such as quantum dots (QDs), allow for a versatile conjugation of various ABs of the same or different kind by several conjugation strategies, and the outstanding photophysical properties of QDs can be very beneficial for both high sensitivity and multiplexed FRET diagnostics. , The tunable PL colors of QDs have been exploited within various multiplexed immunoassay applications, such as multicolor microbead detection or multicolor QD detection on functional microporous membranes . Also, different lanthanide ions, such as Eu 3+ in combination with Sm 3+ , were used for multiplexed immunoassays . However, all of these technologies are heterogeneous, which means that several, often time-consuming, immobilization, washing, incubation, and separation steps are necessary for the complete assay procedure.…”

Section: Introductionmentioning

confidence: 99%

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Nanobodies and Antibodies for Duplexed EGFR/HER2 Immunoassays Using Terbium-to-Quantum Dot FRET

Qiu

Wegner

et al. 2016

Chem. Mater.

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Biosensors based on the combination of semiconductor quantum dots (QDs) and Forster resonance energy transfer (FRET) have demonstrated many advantages for simple, fast, sensitive, and multiplexed diagnostics. However, the implementation of QDs as functional standard materials into homogeneous (single-step) FRET immunoassays has not yet been accomplished, because profound investigations of antibody-conjugation strategies concerning their influence on diagnostic performance for quantifying clinical biomarkers are lacking. Here, we report about a systematic study of size, type, orientation, specificity, nonspecific binding, and cross-reactivity of antibodies conjugated to QDs for single and duplexed EGFR and HER2 immunoassays. Time-gated terbium-to-quantum dot FRET detection on a clinical immunoassay fluorescence plate reader (KRYPTOR) enabled a direct comparison of matuzumab, cetuximab, trastuzumab, and pertuzumab monoclonal antibodies and EgA1, EgB4, 11A4, and 18A12 V H H nanobodies conjugated to 605 and 650 nm emitting QDs. Detection limits of 2.9 ng/mL EGFR, using cetuximab and matuzumab conjugates, and 8.0 ng/mL HER2, using oriented 11A4 and 18A12 conjugates, demonstrated the capability of detecting concentrations well below the clinical cutoff values. Multiplexed assays could quantify EGFR and HER2 at low nanomolar concentrations from the same sample. Our results show that careful optimization of QD-antibody conjugation is a prerequisite to implementing QDs into applied clinical diagnostics.

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Section: ■ Introductionmentioning

confidence: 99%