TY - JOUR A1 - Hildebrandt, Niko A1 - Charbonniere, Loïc J. A1 - Beck, Michael A1 - Ziessel, Raymond F. A1 - Löhmannsröben, Hans-Gerd T1 - Quantum dots as efficient energy acceptors in a time-resolved fluoroimmunoassay Y1 - 2005 SN - 1433-7851 ER - TY - JOUR A1 - Charbonnière, Loic J. A1 - Hildebrandt, Niko T1 - Lanthanide complexes and quantum dots : a bright wedding for resonance energy transfer N2 - In this microreview we describe the principle of Forster resonance energy transfer (FRET) occurring between closely spaced energy-donor and -acceptor molecules. The theoretical treatment is depicted in relation with the data extractable from spectroscopic measurements. We present the specific case of semiconductor nanocrystals (or quantum dots QDs) as energy donors in FRET experiments and a particular emphasis is put on the specific advantages of these fluorophores with regard to both their exceptional photophysical properties and their nanoscopic morphology. In a following section, the special attributes of luminescent lanthanide complexes (LLCs) are outlined with illustrations of properties such as their characteristic emission spectra, long-lived luminescence, and large "Stokes shift". Finally, the successful combination of LLCs and QDs in FRET experiments is demonstrated, showing the unrivaled benefits of this singular marriage, opening doors for energy transfer at very large distances and excellent sensitivity of detection within the frame of time-resolved fluoroimmunoassays. ((C) Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2008). Y1 - 2008 ER - TY - JOUR A1 - Geißler, Daniel A1 - Butlin, Nathaniel G. A1 - Hill, Diana A1 - Löhmannsröben, Hans-Gerd A1 - Hildebrandt, Niko T1 - Multiplexed diagnostics and spectroscopic ruler applications with terbium to quantum dots FRET Y1 - 2008 SN - 1605-7422 ER - TY - JOUR A1 - Kadjane, Pascal A1 - Starck, Matthieu A1 - Camerel, Franck A1 - Hill, Diana A1 - Hildebrandt, Niko A1 - Ziessel, Raymond A1 - Charbonnière, Loïc J. T1 - Divergent approach to a large variety of versatile luminescent lanthanide complexes N2 - Using a regioselective strategy for nucleophilic aromatic substitution on polyfluoropyridines, a nonacoordinating precursor was designed that is adequately suited for complexation of lanthanide cations. Further functionalizations afforded numerous applications for near-IR emission, two-photon absorption spectroscopy, or the formation of luminescent gels. Y1 - 2009 UR - http://pubs.acs.org/journal/inocaj U6 - https://doi.org/10.1021/Ic9001169 SN - 0020-1669 ER - TY - JOUR A1 - Geißler, Daniel A1 - Charbonnière, Loïc J. A1 - Ziessel, Raymond F. A1 - Butlin, Nathaniel G. A1 - Löhmannsröben, Hans-Gerd A1 - Hildebrandt, Niko T1 - Quantum dot biosensors for ultrasensitive multiplexed diagnostics N2 - Time- and color-resolved detection of Foerster resonance energy transfer (FRET) from luminescent terbium complexes to different semiconductor quantum dots results in a fivefold multiplexed bioassay with sub-picomolar detection limits for all five bioanalytes (see picture). The detection of up to five biomarkers occurs with a sensitivity that is 40-240-fold higher than one of the best-established single-analyte reference assays. Y1 - 2010 UR - http://www3.interscience.wiley.com/cgi-bin/jhome/26737/ U6 - https://doi.org/10.1002/anie.200906399 SN - 1433-7851 ER - TY - JOUR A1 - Sellrie, Frank A1 - Beck, Michael A1 - Hildebrandt, Niko A1 - Micheel, Burkhard T1 - A homogeneous time-resolved fluoroimmunoassay (TR-FIA) using antibody mediated luminescence quenching N2 - The determination of low-molecular weight substances (haptens) is demonstrated with a homogeneous time-resolved immunoassay using antibody-induced luminescence quenching. Our novel assay technology uses the newly developed monoclonal antibody (G24-BA9) to quench the luminescence of europium trisbipyridine (EuTBP). We performed a competitive biotin immunoassay including an EuTBP-biotin conjugate, the anti-EuTBP antibody G24-BA9 and streptavidin as assay components. Steric hindrance allows only the binding of either G24-BA9 (to the EuTBP moiety) or streptavidin (to the biotin moiety) to the EuTBP-biotin conjugate. Addition of the analyte biotin resulted in the binding of streptavidin to biotin and a concomitant preferred binding of G24-BA9 to EuTBP-biotin. Since G24-BA9 quenches the luminescence of EuTBP within the conjugate, the luminescence signal could be used to indicate and quantify the presence of free biotin in the system. All experiments were carried out in solution in the presence of 5% serum demonstrating the possibility of using our novel assay for a very fast determination of low molecular weight substances in biological fluids. Y1 - 2010 UR - http://www.rsc.org/Publishing/Journals/AY/Index.asp U6 - https://doi.org/10.1039/C0ay00306a SN - 1759-9660 ER - TY - JOUR A1 - Morgner, Frank A1 - Stufler, Stefan A1 - Geissler, Daniel A1 - Medintz, Igor L. A1 - Algar, W. Russ A1 - Susumu, Kimihiro A1 - Stewart, Michael H. A1 - Blanco-Canosa, Juan B. A1 - Dawson, Philip E. A1 - Hildebrandt, Niko T1 - Terbium to quantum dot FRET Bioconjugates for clinical diagnostics influence of human plasma on optical and assembly properties JF - Sensors N2 - Forster resonance energy transfer (FRET) from luminescent terbium complexes (LTC) as donors to semiconductor quantum dots (QDs) as acceptors allows extraordinary large FRET efficiencies due to the long Forster distances afforded. Moreover, time-gated detection permits an efficient suppression of autofluorescent background leading to sub-picomolar detection limits even within multiplexed detection formats. These characteristics make FRET-systems with LTC and QDs excellent candidates for clinical diagnostics. So far, such proofs of principle for highly sensitive multiplexed biosensing have only been performed under optimized buffer conditions and interactions between real-life clinical media such as human serum or plasma and LTC-QD-FRET-systems have not yet been taken into account. Here we present an extensive spectroscopic analysis of absorption, excitation and emission spectra along with the luminescence decay times of both the single components as well as the assembled FRET-systems in TRIS-buffer, TRIS-buffer with 2% bovine serum albumin, and fresh human plasma. Moreover, we evaluated homogeneous LTC-QD FRET assays in QD conjugates assembled with either the well-known, specific biotin-streptavidin biological interaction or, alternatively, the metal-affinity coordination of histidine to zinc. In the case of conjugates assembled with biotin-streptavidin no significant interference with the optical and binding properties occurs whereas the histidine-zinc system appears to be affected by human plasma. KW - FRET KW - quantum dots KW - terbium KW - luminescence lifetime KW - blood KW - plasma KW - clinical diagnostics KW - biotin KW - streptavidin KW - histidin KW - immunoassay Y1 - 2011 U6 - https://doi.org/10.3390/s111009667 SN - 1424-8220 VL - 11 IS - 10 SP - 9667 EP - 9684 PB - MDPI CY - Basel ER - TY - JOUR A1 - Kupstat, Annette A1 - Kumke, Michael Uwe A1 - Hildebrandt, Niko T1 - Toward sensitive, quantitative point-of-care testing (POCT) of protein markers miniaturization of a homogeneous time-resolved fluoroimmunoassay for prostate-specific antigen detection JF - The analyst : the analytical journal of the Royal Society of Chemistry N2 - Point-of-care testing (POCT) systems which allow for a sensitive, quantitative detection of protein markers are extremely useful for the early detection and therapy progress monitoring of cancer. However, currently commercially available POCT devices are mainly limited to the qualitative detection of protein markers. In this study we demonstrate the successive miniaturization of a sensitive and fast assay for the quantitative detection of prostate-specific antigen (PSA) using a well established and clinically approved homogeneous time-resolved fluoroimmunoassay technology (TRACE (R)) on a commercial plate-reader system (KRYPTOR (R)). Regarding the initial requirements for the development of POCT devices we applied a 30-fold assay volume reduction (150 mu L to 5 mu L) to achieve a reasonable lab-on-a-chip volume and a 24-fold and 120-fold excitation pulse energy reduction to achieve reasonable pulse energies for low-cost miniature excitation sources. Due to highly efficient optimization of key POCT parameters our miniaturized PSA assay achieved a 30% increased sensitivity and a 2-fold improved limit of detection compared to the standard plate-reader method. Our results demonstrate the successful implementation of key parameters for a significant miniaturization and for cost reduction in the clinically approved KRYPTOR (R) platform for protein detection. The technological alterations required are easy-to-implement and can be immediately adapted for more than 30 diagnostic protein markers already available for the KRYPTOR (R) platform. These features strongly recommend our assay format to be utilized in innovative, sensitive, quantitative POCT of protein markers. Y1 - 2011 U6 - https://doi.org/10.1039/c0an00684j SN - 0003-2654 VL - 136 IS - 5 SP - 1029 EP - 1035 PB - Royal Society of Chemistry CY - Cambridge ER - TY - JOUR A1 - Cywinski, Piotr J. A1 - Moro, Artur J. A1 - Ritschel, Thomas A1 - Hildebrandt, Niko A1 - Löhmannsröben, Hans-Gerd T1 - Sensitive and selective fluorescence detection of guanosine nucleotides by nanoparticles conjugated with a naphthyridine receptor JF - Analytical & bioanalytical chemistry N2 - Novel fluorescent nanosensors, based on a naphthyridine receptor, have been developed for the detection of guanosine nucleotides, and both their sensitivity and selectivity to various nucleotides were evaluated. The nanosensors were constructed from polystyrene nanoparticles functionalized by (N-(7-((3-aminophenyl) ethynyl)-1,8-naphthyridin- 2-yl) acetamide) via carbodiimide ester activation. We show that this naphthyridine nanosensor binds guanosine nucleotides preferentially over adenine, cytosine, and thymidine nucleotides. Upon interaction with nucleotides, the fluorescence of the nanosensor is gradually quenched yielding Stern-Volmer constants in the range of 2.1 to 35.9mM(-1). For all the studied quenchers, limits of detection (LOD) and tolerance levels for the nanosensors were also determined. The lowest (3 sigma) LOD was found for guanosine 3',5'-cyclic monophosphate (cGMP) and it was as low as 150 ng/ml. In addition, we demonstrated that the spatial arrangement of bound analytes on the nanosensors' surfaces is what is responsible for their selectivity to different guanosine nucleotides. We found a correlation between the changes of the fluorescence signal and the number of phosphate groups of a nucleotide. Results of molecular modeling and zeta-potential measurements confirm that the arrangement of analytes on the surface provides for the selectivity of the nanosensors. These fluorescent nanosensors have the potential to be applied in multi-analyte, array-based detection platforms, as well as in multiplexed microfluidic systems. KW - Naphthyridine receptor KW - cGMP KW - Base pairing KW - Nucleotide nanosensor KW - Fluorescence spectroscopy Y1 - 2011 U6 - https://doi.org/10.1007/s00216-010-4420-2 SN - 1618-2642 VL - 399 IS - 3 SP - 1215 EP - 1222 PB - Springer CY - Heidelberg ER - TY - JOUR A1 - Hildebrandt, Niko A1 - Geissler, Daniel ED - Zahavy, E ED - Ordentlich, A ED - Yitzhaki, S ED - Shafferman, A T1 - Semiconductor quantum dots as FRET acceptors for multiplexed diagnostics and molecular ruler application JF - Advances in Experimental Medicine and Biology N2 - Applications based on Forster resonance energy transfer (FRET) play an important role for the determination of concentrations and distances within nanometer-scale systems in vitro and in vivo in many fields of biotechnology. Semiconductor nanocrystals (Quantum dots - QDs) possess ideal properties for their application as FRET acceptors when the donors have long excited state lifetimes and when direct excitation of QDs can be efficiently suppressed. Therefore, luminescent terbium complexes (LTCs) with excited state lifetimes of more than 2 ms are ideal FRET donor candidates for QD-acceptors. This chapter will give a short overview of theoretical and practical background of FRET, QDs and LTCs, and present some recent applications of LTC-QD FRET pairs for multiplexed ultra-sensitive in vitro diagnostics and nanometer-resolution molecular distance measurements. KW - Diagnostics KW - FRET KW - Imaging KW - Quantum dots KW - Terbium Y1 - 2012 SN - 978-94-007-2554-6 SN - 978-94-007-2555-3 U6 - https://doi.org/10.1007/978-94-007-2555-3_8 SN - 0065-2598 VL - 733 SP - 75 EP - 86 PB - Springer CY - Dordrecht ER -