Publikationsliste Dr. Nadine Borst

• Originalarbeiten in wissenschaftlichen Fachzeitschriften
• Reviews/Übersichtsartikel in wissenschaftlichen Fachzeitschriften
• Kurzbeiträge
• Vorträge
• Konferenzbeiträge

Originalarbeiten in wissenschaftlichen Fachzeitschriften

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2023

• S. Calabrese, A. M. Markl, M. Neugebauer, S. J. Krauth, N. Borst, F. von Stetten, M. Lehnert
  Reporter emission multiplexing in digital PCRs (REM-dPCRs): direct quantification of multiple target sequences per detection channel by population specific reporters
  2023 Analyst, Band: 148, Seiten: 5243 - 5254

2022

• E. Kipf, F. Schlenker, N. Borst, M. Fillies, R. Kirschner-Schwabe, R. Zengerle, C. Eckert, F. von Stetten, M. Lehnert
  Advanced minimal residual disease monitoring for acute lymphoblastic leukemia with multiplex mediator probe PCR
  2022 J. Mol. Diag, Band: 24, Nummer: 1, Seiten: 57 - 68
• Y.-T. Kao, S. Calabrese, N. Borst, M. Lehnert, Y.-K. Lai, F. Schlenker, P. Juelg, R. Zengerle, P. Garstecki, F. von Stetten
  Microfluidic One-Pot Digital Droplet FISH Using LNA/DNA Molecular Beacons for Bacteria Detection and Absolute Quantification
  2022 Biosensors, Band: 12, Nummer: 4, Seite: 237

2021

• F. Schlenker, E. Kipf, N. Borst, N. Paust, R. Zengerle, F. von Stetten, P. Juelg, T. Hutzenlaub
  Centrifugal Microfluidic Integration of 4-Plex ddPCR Demonstrated by the Quantification of Cancer-Associated Point Mutations
  2021 Processes, Band: 2021, Nummer: 9, Ergänzungsband: 97
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  Kurzfassung

  We present the centrifugal microfluidic implementation of a four-plex digital droplet polymerase chain reaction (ddPCR). The platform features 12 identical ddPCR units on a LabDisk cartridge, each capable of generating droplets with a diameter of 82.7 +/- 9 µm. By investigating different oil–surfactant concentrations, we identified a robust process for droplet generation and stabilization. We observed high droplet stability during thermocycling and endpoint fluorescence imaging, as is required for ddPCRs. Furthermore, we introduce an automated process for fourcolor fluorescence imaging using a commercial cell analysis microscope, including a customized software pipeline for ddPCR image evaluation. The applicability of ddPCRs is demonstrated by the quantification of three cancer-associated KRAS point mutations (G12D, G12V and G12A) in a diagnostically relevant wild type DNA background. The four-plex assay showed high sensitivity (3.5–35 mutant DNA copies in 15,000 wild type DNA copies) and linear performance (R2 = 0.99) across all targets in the LabDisk.
• L. Becherer, J. F. Hess, S. Frischmann, M. Bakheit, H. Nitschko, S. Stinco, F. Zitz, H. Hofer, G. Porro, F. Hausladen, K. Stock, D. Drossart, H. Wurm, H. Kuhn, D. Huber, T. Hutzenlaub, N. Paust, M. Keller, O. Strohmeier, S. Wadle, N. Borst, R. Zengerle, F. von Stetten
  Point-of-Care System for HTLV-1 Proviral Load Quantification by Digital Mediator Displacement LAMP
  2021 Micromachines, Band: 12, Nummer: 2, Seite: 159
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  Kurzfassung

  This paper presents a universal point-of-care system for fully automated quantification of human T-cell lymphotropic virus type 1 (HTLV-1) proviral load, including genomic RNA, based on digital reverse RNA transcription and c-DNA amplification by MD LAMP (mediator displacement loop-mediated isothermal amplification). A disposable microfluidic LabDisk with pre-stored reagents performs automated nucleic acid extraction, reaction setup, emulsification, reverse transcription, digital DNA amplification, and quantitative fluorogenic endpoint detection with universal reporter molecules. Automated nucleic acid extraction from a suspension of HTLV-1-infected CD4+ Tlymphocytes (MT-2 cells) yielded 8 +/- 7 viral nucleic acid copies per MT-2 cell, very similar to the manual reference extraction (7 +/- 2 nucleic acid copies). Fully automated sample processing from whole blood spiked with MT-2 cells showed a comparable result of 7 +/- 3 copies per MT-2 cell after a run time of two hours and 10 min.
• F. Schlenker, E. Kipf, M. Deuter, I. Höffkes, M. Lehnert, R. Zengerle, F von Stetten, F. Scherer, J. Wehrle, N. von Bubnoff, P. Juelg, T. Hutzenlaub, N. Borst
  Stringent Base Specific and Optimization-Free Multiplex Mediator Probe ddPCR for the Quantification of Point Mutations in Circulating Tumor DNA
  2021 cancers, Band: 13, Nummer: 22, Seite: 5742
• F. Schlenker, E. Kipf, N. Borst, T. Hutzenlaub, R. Zengerle, F. von Stetten, P. Juelg
  Virtual Fluorescence Color Channels by Selective Photobleaching in Digital PCR Applied to the Quantification of KRAS Point Mutations
  2021 Analytical Chemistry, Band: 93, Nummer: 30, Seiten: 10538 - 10545
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  Kurzfassung

  Multiplexing of analyses is essential to reduce sample and reagent consumption in applications with large target panels. In applications such as cancer diagnostics, the required degree of multiplexing often exceeds the number of available fluorescence channels in polymerase chain reaction (PCR) devices. The combination of photobleaching-sensitive and photobleaching-resistant fluorophores of the same color can boost the degree of multiplexing by a factor of 2 per channel. The only additional hardware required to create virtual fluorescence color channels is a low-cost light-emitting diode (LED) setup for selective photobleaching. Here, we present an assay concept for fluorescence color multiplexing in up to 10 channels (five standard channels plus five virtual channels) using the mediator probe PCR with universal reporter (UR) fluorogenic oligonucleotides. We evaluate the photobleaching characteristic of 21 URs, which cover the whole spectral range from blue to crimson. This comprehensive UR data set is employed to demonstrate the use of three virtual channels in addition to the three standard channels of a commercial dPCR device (blue, green, and red) targeting cancer-associated point mutations (KRAS G12D and G12V). Moreover, a LOD (limit of detection) analysis of this assay confirms the high sensitivity of the multiplexing method (KRAS G12D: 16 DNA copies/reaction in the standard red channel and KRAS G12V: nine DNA copies/reaction in the virtual red channel). Based on the presented data set, optimal fluorogenic reporter combinations can be easily selected for the application-specific creation of virtual channels, enabling a high degree of multiplexing at low optical and technical effort.

2020

• M. Trotter, D. Juric, Z. Bagherian, N. Borst, K. Gläser, T. Meissner, F. von Stetten, A. Zimmermann
  Inkjet-Printing of Nanoparticle Gold and Silver Ink on Cyclic Olefin Copolymer for DNA-Sensing Applications
  2020 Sensors, Band: 20, Nummer: 5, Seite: 1333
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  Kurzfassung

  Inkjet technology as a maskless, direct-writing technology offers the potential for structured deposition of functional materials for the realization of electrodes for, e.g., sensing applications. In this work, electrodes were realized by inkjet-printing of commercial nanoparticle gold ink on planar substrates and, for the first time, onto the 2.5D surfaces of a 0.5 mm-deep microfluidic chamber produced in cyclic olefin copolymer (COC). The challenges of a poor wetting behavior and a low process temperature of the COC used were solved by a pretreatment with oxygen plasma and the combination of thermal (130 °C for 1 h) and photonic (955 mJ/cm²) steps for sintering. By performing the photonic curing, the resistance could be reduced by about 50% to 22.7 μΩ cm. The printed gold structures were mechanically stable (optimal cross-cut value) and porous (roughness factors between 8.6 and 24.4 for 3 and 9 inkjet-printed layers, respectively). Thiolated DNA probes were immobilized throughout the porous structure without the necessity of a surface activation step. Hybridization of labeled DNA probes resulted in specific signals comparable to signals on commercial screen-printed electrodes and could be reproduced after regeneration. The process described may facilitate the integration of electrodes in 2.5D lab-on-a-chip systems.
• L. Becherer, N. Borst, M. Bakheit, S. Frischmann, R. Zengerle, F. von Stetten
  Loop-mediated isothermal amplification (LAMP) – review and classification of methods for sequence-specific detection
  2020 Anal Methods-uk, Band: 12, Seiten: 717 - 746
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  Kurzfassung

  In the course of the last 20 years, isothermal nucleic acid amplification tests have emerged as an important diagnostic tool, not only for clinical applications, but also for food quality control and environmental monitoring. Loop-mediated isothermal amplification (LAMP) is well known for its robust and highly sensitive and specific amplification of target DNA, which is achieved by utilizing up to six primers. Moreover, LAMP excels through its isothermal and energy efficient amplification requirements, rendering it a prime candidate for low-cost diagnostics and analysis at the point of need. Recently, methods for sequence-specific detection have gained more importance because, unlike sequence-independent detection methods, they are highly specific towards the target DNA. In the last 13 years, a variety of sequence-specific methods have emerged, based on a very diverse range of sensing techniques, including optical, magnetic, piezoelectric, electrochemical and magnetoresistive sensing. To give structure to the diverse multitude of sequence-specific methods, we created a systematic classification and provide a critical comparative evaluation according to a catalogue of criteria (analytical performance, multiplexing, quantification and instrumental requirements). Fluorescence-based detection, making up half of the methods, can be processed on open platforms and satisfies all the criteria listed before. Instrumental requirements are discussed in terms of complexity, portability and fluidic cartridges. In addition, the technological readiness level and the kind of platform (open versus method-tailored) are evaluated, the latter playing an important role in the miniaturization and automation of operational process steps. We also observe an increase in the use of smartphone-integrated sensors to improve LAMP-based point-of-need testing. In summary, recent developments in methods for the sequence-specific detection of LAMP demonstrate high potential for many future applications.
• L. Becherer, S. Knauf, M. Marks, S. Lueert, S. Frischmann, N. Borst, F. von Stetten, S. Bieb, Y. Adu-Sarkodie, K. Asiedu, O. Mitjà, M. Bakheit
  Multiplex Mediator Displacement Loop-Mediated Isothermal Amplification for Detection of Treponema pallidum and Haemophilus ducreyi
  2020 Emerg Infect Dis, Band: 26, Seiten: 282 - 288
• M. Schulz, S. Calabrese, F. Hausladen, H. Wurm, D. Drossart, K. Stock, A. M. Sobieraj, F. Eichenseher, M. J. Loessner, M. Schmelcher, A. Gerhardts, U. Goetz, M. Handel, A. Serr, G. Haecker, J. Li, M. Specht, P. Koch, M. Meyer, P. Tepper, R. Rother, M. Jehle, S. Wadle, R. Zengerle, F. von Stetten, N. Paust, N. Borst
  Point-of-care testing system for digital single cell detection of MRSA directly from nasal swabs
  2020 Lab Chip, Band: 20, Seiten: 2549 - 2561
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  Kurzfassung

  We present an automated point-of-care testing (POCT) system for rapid detection of species- and resistance markers in methicillin-resistant Staphylococcus aureus (MRSA) at the level of single cells, directly from nasal swab samples. Our novel system allows clear differentiation between MRSA, methicillin-sensitive S. aureus (MSSA) and methicillin-resistant coagulase-negative staphylococci (MR-CoNS), which is not the case for currently used real-time quantitative PCR based systems. On top, the novel approach outcompetes the culture-based methods in terms of its short time-to-result (1 h vs. up to 60 h) and reduces manual labor. The walk-away test is fully automated on the centrifugal microfluidic LabDisk platform. The LabDisk cartridge comprises the unit operations swab-uptake, reagent pre-storage, distribution of the sample into 20 000 droplets, specific enzymatic lysis of Staphylococcus spp. and recombinase polymerase amplification (RPA) of species (vicK) – and resistance (mecA) -markers. LabDisk actuation, incubation and multi-channel fluorescence detection is demonstrated with a clinical isolate and spiked nasal swab samples down to a limit of detection (LOD) of 3 ± 0.3 CFU μl−1 for MRSA. The novel approach of the digital single cell detection is suggested to improve hospital admission screening, timely decision making, and goal-oriented antibiotic therapy. The implementation of a higher degree of multiplexing is required to translate the results into clinical practice.
• M. Schulz, S. Probst, S. Calabrese, A. Homann, N. Borst, M. Weiss, F. von Stetten, R. Zengerle, N. Paust
  Versatile Tool for Droplet Generation in Standard Reaction Tubes by Centrifugal Step Emulsification
  2020 Molecules, Band: 25, Nummer: 8, Seite: 1914
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  Kurzfassung

  We present a versatile tool for the generation of monodisperse water-in-fluorinated-oil droplets in standard reaction tubes by centrifugal step emulsification. The microfluidic cartridge is designed as an insert into a standard 2 mL reaction tube and can be processed in standard laboratory centrifuges. It allows for droplet generation and subsequent transfer for any downstream analysis or further use, does not need any specialized device, and manufacturing is simple because it consists of two parts only: A structured substrate and a sealing foil. The design of the structured substrate is compatible to injection molding to allow manufacturing at large scale. Droplets are generated in fluorinated oil and collected in the reaction tube for subsequent analysis. For sample sizes up to 100 µL with a viscosity range of 1 mPa·s–4 mPa·s, we demonstrate stable droplet generation and transfer of more than 6 × 105 monodisperse droplets (droplet diameter 66 µm ± 3 µm, CV ≤ 4%) in less than 10 min. With two application examples, a digital droplet polymerase chain reaction (ddPCR) and digital droplet loop mediated isothermal amplification (ddLAMP), we demonstrate the compatibility of the droplet production for two main amplification techniques. Both applications show a high degree of linearity (ddPCR: R2 ≥ 0.994; ddLAMP: R2 ≥ 0.998), which demonstrates that the cartridge and the droplet generation method do not compromise assay performance.

2018

• M. Lehnert, E. Kipf, F. Schlenker, N. Borst, R. Zengerle, F. von Stetten
  Fluorescence signal-to-noise optimisation for real-time PCR using universal reporter oligonucleotides
  2018 Anal Methods-uk
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  Kurzfassung

  In this study we optimised the fluorescence signal generation of contact quenched universal reporter oligonucleotides. These are used as secondary probes in real-time Mediator Probe PCR to detect the sequence-specific cleavage of label-free primary mediator probes. Since the fluorescence signal generation of a universal reporter is not influenced by the target DNA sequence, optimisation of the fluorescence signal-to-noise ratio will improve the performance of all Mediator Probe PCRs that are based on this type of universal reporter. To determine the critical factors influencing signal-to-noise optimisation, we systematically analysed four parameters. These parameters were type of fluorophore, type of quencher molecule, intramolecular orientation of both residuals, and the number of quencher labels. In total, more than 30 different fluorogenic universal reporter structures were analysed, covering the whole fluorescence spectrum from green to crimson. From our results, we deduced a novel set of guidelines for signal-to-noise optimisation in the design of contact quenched, fluorogenic universal reporter oligonucleotides. We confirmed these guidelines in a different thermocycler, and by designing a second set of universal reporters, which were used for multiplex real-time PCR quantification of acute lymphoblastic leukaemia marker sequences. This optimised biplex Mediator Probe PCR showed an improved performance under clinical conditions, with a 10 times higher resolution regarding the limit of quantification. In addition to Mediator Probe PCR, these guidelines may also prove useful in signal-to-noise optimisation of other fluorescence-based assays where contact quenched oligonucleotides or secondary reporter molecules are used.
• L. Becherer, M. Bakheit, S. Frischmann, S. Stinco, N. Borst, R. Zengerle, F. von Stetten
  Simplified real-time multiplex detection of loop-mediated isothermal amplification (LAMP) using novel mediator displacement probes with universal reporters
  2018 Anal Chem, Band: 90, Seiten: 4741 - 4748
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  Kurzfassung

  A variety of real-time detection techniques for LAMP based on the change in fluorescence intensity during DNA amplification enable simultaneous detection of multiple targets. However these techniques depend on fluorogenic probes containing target-specific sequences. That complicates the adaption to different targets leading to time-consuming assay optimization. Here, we present the first universal real-time detection technique for multiplex LAMP. The novel approach allows simple assay design and is easy to implement for various targets. The innovation features a mediator displacement probe and a universal reporter. During amplification of target DNA the mediator is displaced from the mediator displacement probe. Then it hybridizes to the reporter generating a fluorescence signal. The novel Mediator Displacement (MD) detection was validated against state-of-the-art molecular beacon (MB) detection by means of a HIV-1 RT-LAMP: MD surpassed MB detection by accelerated probe design (MD: 10 min, MB: 3-4 h), shorter times to positive (MD 4.1±0.1 min shorter than MB, n = 36), improved signal to noise fluorescence ratio (MD: 5.9±0.4, MB: 2.7±0.4; n = 15) and showed equally good or better analytical performance parameters. The usability of one universal mediator-reporter set in different multiplex assays was successfully demonstrated for a biplex RT-LAMP of HIV-1 & HTLV-1 and a biplex LAMP of Haemophilus ducreyi & Treponema pallidum, both showing good correlation between target concentration and time to positive. Due to its simple implementation it is suggested to extend the use of the universal mediator-reporter sets to the detection of various other diagnostic panels.

2017

• N. Borst, F. Schuler, S. Wadle, M. Schulz, M. Specht, J. Li, L. Becherer, M. Trotter, A. B. Rodríguez-Martínez, N. Paust, R. Zengerle, F. von Stetten
  A technology platform for digital nucleic acid diagnostics at the point of care
  2017 Laboratoriumsmedizin, Band: 41, Nummer: 5
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  Kurzfassung

  The combination of digital amplification and centrifugal microfluidics can enable quantitative and fast diagnostics at the point of care (PoC). The new unit operation of centrifugal step emulsification allows high throughput droplet generation. Different methods for digital nucleic acid analysis, including PCR, recombinase polymerase amplification (RPA) and loop mediated isothermal amplification (LAMP), have already been demonstrated. Our novel approach of integrated sample-to-answer analysis is introduced, and examples for the detection of HIV and single cell analysis of antibiotic resistant bacteria are presented. Next to these LabDisk based systems, a microfluidic cartridge termed DropChip allows for digital amplification using only commercially available laboratory devices.

Reviews/Übersichtsartikel in wissenschaftlichen Fachzeitschriften

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2020

• M. Trotter, N. Borst, R. Thewes, F. von Stetten
  Electrochemical DNA sensing – Principles, commercial systems, and applications
  2020 Biosens Bioelectron, Band: 154, Seite: 112069
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  Kurzfassung

  Driven by the vision of robust and portable, yet sensitive DNA detection systems for point-of-need applications, the development of electrochemical DNA sensing principles has been of high interest. Many different principles have been developed and these are regularly reviewed. However, the maturity of electrochemical principles and their ability to produce competitive real-world applications is rarely assessed. In this review, general electrochemical DNA sensing principles are briefly introduced and categorized into heterogeneous vs. homogeneous approaches, and then the subcategories label-free vs. labeled and reagent-less vs. reagent-dependent principles. We then focus on reviewing the electrochemical sensing principles implemented in DNA detection systems, which are commercially available or close to market entry, considering the complete analysis process, automation and the field of application. This allows us to outline and discuss which principles have proved suitable for which kinds of applications, as well as the stage of integration and automation. Examples from all the identified categories of electrochemical DNA sensing principles have found application in commercial detection systems or advanced prototypes. Various applications have already been demonstrated, ranging from on-site skin care testing, to food safety to the most frequent in vitro diagnostic tests, partially conducted in automated sample-to-answer devices. Our review is intended to enable researchers in areas related to electrochemistry, biochemistry or microfluidics to assess the commercial state of the art of electrochemical nucleic acid testing, and the interdisciplinary challenges for further improvements.
  
  
   
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Kurzbeiträge

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2017

• N. Borst, F. von Stetten
  Eine Technologieplattform für die digitale Nukleinsäureanalytik
  2017 Management & Krankenhaus, Band: 2017-04, Seite: 37

Vorträge

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2017

• F. Schuler, S. Wadle, N. Borst, M. Schulz, L. Becherer, J. Li, M. Specht, T. Hutzenlaub, N. Paust, R. Zengerle, F. von Stetten
  A Technology Platform for Digital Nucleic Acid Diagnostics at the Point of Care (invited talk)
  2017 3. Münchner point-of-care testing symposium, Deutsche Gesellschaft für Klinische Chemie und Laboratoriumsmedizin (DGKL), München, 13.-15. 3. 2017
• F. Schuler, N. Borst, S. Wadle, M. Schulz, M. Specht, J. Li, L. Becherer, T. Hutzenlaub, N. Paust, R. Zengerle, F. von Stetten
  Centrifugal Step Emulsification Allows Miniaturized Digital Droplet-RPA,-LAMP and -PCR on the Centrifugal Microfluidic Platform (invited talk)
  2017 9th Annual lab-on-a-chip & microfluidics Conference, Selectbio, Munich, 10.-11.5. 2017

Konferenzbeiträge

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2023

• H. Gmoser, T. Groß, M. Ballweg, A. Lux, C. Jeney, M. Lehnert, F. v. Stetten, N. Borst
  Isothermal Protein Interaction Coupling (PICO) Assay towards One-Step Library Prep and Nanopore Read-out
  2023 Black Forest Nanopore Meeting, Freiburg, 06.-09.11.2023
• N. Borst
  Neue Technologien für die molekulare Diagnostik zuhause
  2023 HomeDiagnostix Congress, Konstanz, July 5-7, 2023

2022

• M. Trotter, A. Schreiber, D. Kleinknecht, N. Borst, M. Kuderer, K. Gläser, F. von Stetten
  Mobile elektrochemische Plattform zur molekularen Diagnostik in der häuslichen Pflege
  2022 5. Müchner Poit-of-Care Testing Symposium, Munich, September,27-29, 2022

2021

• M. Trotter, D. Juric, Z. Bagherian, N. Borst, K. Gläser, F. von Stetten, A. Zimmermann
  Integration of nanoporous gold electrodes in microfluidic chambers by inkjet-printing for biosensing applications
  2021 MST-Kongress, Ludwigsburg, 08.-10.11.2021
• Y.-T. Kao, S. Calabrese, N. Borst, M. Lehnert, Y.-K. Lai, F. Schlenker, R. Zengerle, P. Garstecki, F. von Stetten
  One-step amplification-free bacteria detection by optimizes LNA/DNA molecular beacons in droplets
  2021 MicroTAS 2021, Palm Springs/USA, 10.-14.10.2021, online
• L. Becherer, J.F. Hess, S. Frischmann, M. Bakheit, H. Nitschko, S. Stinco, F. Zitz, H. Hofer, G. Porro, F. Hausladen, K. Stock, D. Drossart, H. Wurm, H. Kuhn, D. Huber, T. Hutzenlaub, N. Paust, M. Keller, O. Strohmeier, S. Wadle, N. Borst, R. Zengerle, F. von Stetten
  Point-of-Care System for HTLV-1 Proviral Load Quantification by Digital Mediator Displacement LAMP
  2021 MST-Kongress, Ludwigsburg, 08.-10.11.2021
• T. Härpfer, L. Becherer, M. Bakheit, S. Frischmann, S. Stinco, N. Borst, R. Zengerle, F. von Stetten
  Universal multiplex mediator displacement LAMP Nr. P3.062 analytical and clinical validation on the example of HIV, Haemophilus ducreyi and Treponema pallidum
  2021 Biosensors, , Haeundae-gu, South Korea (online) , 26. – 29.07.2021

2020

• F. Schlenker, E. Kipf, N. Borst, T. Hutzenlaub, N. Paust, R. Zengerle, F. von Stetten, P. Juelg
  Centrifugal microfluidic 4 Plex digital droplet PCR for quantification of circulating tumor DNA
  2020 MicroTAS 2020, 04.-09.10.2020, virtual
• F. Schlenker, E. Kipf, M. Deuter, I. Hoeffkes, N. Borst, T. Hutzenlaub, R. Zengerle, F. von Stetten, N. von Bubnoff, P. Juelg
  Low optimization effort for highly sensitive assays for point mutation quantification in CTDNA: the Mediator Probe digital PCR
  2020 Liquid Biopsy Symposium, 30.10.2020, virtuell

2019

• M. Lehnert, E. Kipf, F. Schlenker, R. Zengerle, N. Borst, F. von Stetten
  Multiplex Mediator Probe Real-Time PCR - Optimization and Guideline Development through Systematic Characterisation of Label Free Mediator Probes and Fluorogenic Universal Reporters
  2019 9th Gene Quantification Event qPCR dPCR & NGS 2019, 18 - 22 March 2019, in Freising-Weihenstephan, Germany
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  Kurzfassung

  Mediator Probe PCR is a powerful and robust real-time PCR technology for multiplex DNA detection and quantification. It uses label free mediator probes, for molecular detection of nucleic acids during DNA amplification, in combination with fluorogenic universal reporters for signal generation. During PCR, target sequence specific mediator probes are cleaved by the polymerase and a generic sequence, the mediator, is set free. In the second step the mediator binds to the universal reporter, where it is extended by the polymerase. This generates a strong fluorescence signal increase. Due to the separation of DNA detection and signal generation many advantages arise. Mediator probes are not limited in their design by properties of the target sequence and a standard set of highly optimised fluorogenic universal reporters can be used for multiplex Mediator Probe PCR, right from the start.1 In the last years Mediator Probe PCR evolved from an innovative new method to an optimised and robust multiplexing technology. This was achieved by systematic characterisation of its molecular processes, which again was advantaged by the separation of DNA detection and signal generation. A DoE approach was used for the optimisation of Mediator Probes, focusing on their binding strengths.2 In parallel, a set of universal reporters with improved signal-to-noise ratios was established by successive testing over 40 molecular structures with different fluorophore-quencher labels and configurations.1 As a result, distinct guidelines exist, which enable fast adaption of new DNA targets and facilitate multiplex Mediator Probe PCR design. The capability of the technology was shown by highly sensitive, precise and specific multiplex Mediator Probe real-time PCRs in different areas of molecular diagnostics. These fields include monitoring of oncological disease, detection of pathogens or analysis of food samples.1,3
• F. Schlenker, E. Kipf, S. Jenne, N. Borst, J. Lüddecke, K. Dormanns, T. Hutzenlaub, J. Steinert, N. Paust, R. Zengerle, F. von Stetten, P. Juelg
  Doubling the order of color-multiplexing by photobleaching in Mediator Probe droplet digital PCR
  2019 EACR-ESMO Joint Conference on Liquid Biopsies, Bergamo/Italien, 15. – 17. 05. 2019

2018

• Z. Bagheryan, M. Trotter, L. Becherer, N. Borst, F. von Stetten
  A universal approach for electrochemically DNA detection based on mediator displacement LAMP
  2018 9th Euro Biosensors & Bioelectronics Congress November 29-30, 2018 Dublin, Ireland
• L. Becherer, M. Bakheit, S. Frischmann, S. Stinco, N. Borst, R. Zengerle, F. von Stetten
  Fluorogenic real-time detection of loop-mediated isothermal amplification by novel mediator displacement probes demonstrated for HIV-1 and HTLV-1
  2018 International Biotech Innovation Days, 23 - 25 May 2018 / Senftenberg, Germany
• M. Schulz, N. Borst, M. Specht, S. Calabrese, F. von Stetten, R. Zengerle, N. Paust
  From nasal swab to digital answer: unit operations for antibiotic resistance screening on a single cell level
  2018 MicroTAS 2018, 11. -15. November 2018, Kaohsiung / Taiwan
• L. Becherer, M. Schulz, H. Kuhn, M. Bakheit, S. Frischmann, F. Zitz, N. Borst, R. Zengerle, F. von Stetten
  HIV-1 and HTLV-1 multiplex detection by digital mediator displacement LAMP
  2018 MicroTAS 2018, 11. -15. November 2018, Kaohsiung / Taiwan
• M. Trotter, N. Borst, F. von Stetten
  Mediator probes for electrochemical DNA detection: Universal electrode functionalization for specific detection of different targets
  2018 9th Euro Biosensors & Bioelectronics Congress November 29-30, 2018 Dublin, Ireland
• L. Becherer, N. Borst, R. Zengerle, F. von Stetten
  Simultaneous detection of HIV and HTLV by mediator displacement loop-mediated isothermal amplification
  2018 6th World Congress on Control and Prevention of HIV/AIDS, STDs & STIs, Zürich/Switzerland, 27. – 28.08.2018
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  Kurzfassung

  6th World Congress on Control and Prevention of HIV/AIDS, STDs & STIs, Zürich/Switzerland, 27. – 28.08.2018

2017

• F. Schuler, N. Borst, S. Wadle, M. Schulz, M. Specht, J. Li, L. Becherer, T. Hutzenlaub, N. Paust, R. Zengerle, F. v. Stetten
  Eine Technologieplattform für die digitale Nukleinsäurediagnostik vor Ort (A Technology Platform for Digital Nucleic Acid Diagnostics at the Point of Care)
  2017 3. Münchner Point-of-Care-Testing Symposium der DGKL (Deutsche Gesellschaft für Klinische Chemie und Laboratoriumsmedizin e.V.) 13. – 15. März 2017, München
• N. Borst
  Innovative Diagnostikplattform zum Nachweis antibiotikaresistenter Erreger im Krankenhaus
  2017 Innovation Forum Medizintechnik 2017, Tuttlingen, 19.10.2017
• N. Borst, S. Wadle, K. Stock, M. Röder, E. Held-Föhn, A. Heiß, U. Götz, F. von Stetten
  Innovative Diagnostikplattform zur Zuordnung von Krankenhauskeimen und deren Antibiotikaresistenzen
  2017 microTEC Südwest Clusterkonferenz 2017, 27.-28. März 2017, Freiburg
• N. Borst, M. Specht, M. Schulz, N. Paust, J. Li, S. Wadle, U. Götz, E. Held-Föhn, A. Heiß, F. Hausladen, R. Mader, K. Stock, M. Röder, A. Serr, G. Häcker, R. Zengerle, F. von Stetten
  Integriertes System zur simultanen Detektion von Krankenhauskeimen und deren Antibiotikaresistenzen auf Einzelzellebene
  2017 MST Kongress, München, 23. - 25.10.2017
  » Kurzfassung anzeigen« Kurzfassung verbergen
  Kurzfassung

  Bei Krankenhausinfektionen mit mehreren beteiligten Erregern ist die Zuordnung von Antibiotikaresistenzen zu den Erregern besonders wichtig um eine spezifische Therapie zu ermöglichen. Ein Projektkonsortium aus 6 Instituten der Innovationsallianz Baden Württemberg entwickelt im Projekt IDAK ein integriertes System das es künftig ermöglichen soll auf Einzelzellebene simultan die Spezies und die Resistenz aller Erreger in einer Probe zu bestimmen. Wir berichten von den aktuellen Forschungsarbeiten und ersten Ergebnissen. Diese umfassen: Entwicklung eines mikrofluidischen Testträgers zur Integration aller Prozessschritte; Überführung einzelner Bakterien eines Nasalabstriches in Tröpfchen; Bakterienlyse; digitaler multiplex-DNA Nachweis auf Basis einer isothermen Amplifikation; Multispektrale Fluoreszenzdetektion der Genotypen.
• A. B. Rodríguez-Martínez, N. Borst, E. Sarasola-Díez, M. Agirregabiria, J. M. Ruano-López, T. Codd, K. R. Ward, J. Bekacz, M. Gaboyard, M. J. García-Barcina
  Novel ddPCR approach for NIPT rhesus genotyping
  2017 European Human Genetics Conference, May 27–30, 2017, Copenhagen / Dänemark