NextGen-O2k

NextGen-O2k

The revolutionary all-in-one instrument to conquer mitochondrial disease

NextGen-O2k: Horizon 2020 Framework Programme

EIC-SMEInst-2018-2020 — SME instrument phase 2, Grant Agreement No. 859770, Oroboros Instruments
Budget: € 2.35 Million
Duration: 24 months, extended additional 5 months
Application: 2019-01-09; Pitch in Brussels: 2019-02-15; Granted: 2019-03-05; Start: 2019-06-01

NextGen-O2k publications, preprints, and abstracts

Link	Reference	View	Year
MiPNet24.12 NextGen-O2k: Q-Module	NextGen-O2k: Q-Module manual	» Versions	2021-06-24
Went 2021 MitoFit PB	Went N, Di Marcello M, Gnaiger E (2021) Oxygen dependence of photosynthesis and light-enhanced dark respiration studied by High-Resolution PhotoRespirometry. MitoFit Preprints 2021.5. doi:10.26124/mitofit:2021-0005	Oxygen dependence of photosynthesis and light-enhanced dark respiration studied by High-Resolution PhotoRespirometry	2021-04-30
Ravasz 2019 Abstract IOC141	Ravasz D, Bui D, Kitayev A, Greenwood B, Hill C, Komlodi T, Doerrier C, Ozohanics O, Moore AL, Gnaiger E, Kiebish M, Kolev K, Seyfried TN, Willis WT, Narain N, Adam-Vizi V, Chinopoulos C (2019) Endogenous quinones sustain a moderate NADH oxidation by Complex I during anoxia. Mitochondr Physiol Network 24.02.
Komlodi 2021 MitoFit Q	Komlodi T, Cardoso LHD, Doerrier C, Moore AL, Rich PR, Gnaiger E (2021) Coupling and pathway control of coenzyme Q redox state and respiration in isolated mitochondria. MitoFit Preprints 2021.2.v2. doi:10.26124/mitofit:2021-0002.v2	Coupling and pathway control of coenzyme Q redox state and respiration in isolated mitochondria	2021-05-06
Huete-Ortega 2020 MitoFit Preprint Arch EA	Huete-Ortega M, Di Marcello M, Iglesias-Gonzalez J, Gnaiger E (2020) High-resolution respirometry for chloroplast and mitochondrial bioenergetics in Chlamydomonas reinhardtii ― towards biotechnology exploitations. MitoFit Preprint Arch doi:10.26124/mitofit:ea20.algaeurope.0001.	*High-resolution respirometry for chloroplast and mitochondrial bioenergetics in Chlamydomonas reinhardtii* ― towards biotechnology exploitations**	2020-11-30
Huete-Ortega M 2021 MitoFit Photosynthesis protocols	Huete-Ortega M, Di Marcello M, Doerrier C, Gnaiger E (2021) Substrate-uncoupler-inhibitor-titration protocols for photosynthesis in Chlamydomonas reinhardtii. MitoFit Preprints 2021 (in prep).
Doerrier 2021 MitoFit NADH	Doerrier C, Gnaiger E (2021) Simultaneous measurement of oxygen consumption and NADH redox states as a metabolic marker in pathophysiological conditions. (in prep)
Copsey 2021 Sci Rep	Copsey AC, Barsottini MRO, May B, Xu F, Albury MS, Young L, Moore AL (2021) Kinetic characterisation and inhibitor sensitivity of Candida albicans and Candida auris recombinant AOX expressed in a self-assembled proteoliposome system. Sci Rep 11:14748.	PMID: 34285303 Open Access	2021
Cardoso 2021 BEC MgG	Cardoso LHD, Doerrier C, Gnaiger E (2021) Magnesium Green for fluorometric measurement of ATP production does not interfere with mitochondrial respiration. Bioenerg Commun 2021.1. doi:10.26124/bec:2021-0001	Published online: 2021-Jun-30	2021-006-30
Cardoso 2021 MitoFit MgG	Cardoso LHD, Doerrier C, Gnaiger E (2021) Magnesium Green for fluorometric measurement of ATP production does not interfere with mitochondrial respiration. MitoFit Preprints 2021.1. doi:10.26124/mitofit:2021-0001	Magnesium Green for fluorometric measurement of ATP production does not interfere with mitochondrial respiration	2021-01-29

About NextGen-O2k

Oroboros - as a driving force in mitochondrial physiology - extends the analytical and diagnostic power of high-resolution respirometry by integration of NADH- and Q-redox monitoring in the NextGen-O2k. We aim at establishing the Oroboros quality control management for dissemination to our worldwide O2k-Network laboratories. This will become an effective contribution to address the acute reproducibility crisis of scientific investigation. In the spirit of Open Science and global networking, we will enable data sharing across projects and institutions in an Open Access database on mitochondrial physiology and pathology, to resolve the inflation crisis and ultimately the value-impact crisis of present academic publication. This will support key developments in mitochondrial medicine. In addition, we expand our business to algal biotechnology and ecology with the photobiology module of the NextGen-O2k, widening our focus from medicine to environment and climate.

- a partner in the Oroboros Ecosystem

Aims

Form the O2k to the NextGen-O2k

Between 1992 and 1994, Oroboros Instruments released its core technology for high-resolution respirometry (HRR), the Oxygraph-2k or O2k, for real-time monitoring of oxygen consumption of mitochondria, tissues, and living cells. Dissolved oxygen is measured using high-resolution polarographic oxygen sensors. Its main advantages are the unique signal stability and resolution of oxygen. HRR is the core technology present in every single of the devices and it will remain the basis of the NextGen-O2k.

In 2012, the updated version of the O2k was launched, the O2k-FluoRespirometer. The integrated Smart Fluo-Sensors combine fluorescence excitation and detection of defined wavelengths that in combination with fluorophores allow for the measurement of additional mitochondrial parameters, such as membrane potential, H₂O₂ production, calcium concentration, and ATP production.

The unique software DatLab has been developed for data acquisition and analysis. DatLab allows the researcher to perform calibrations (including automatic barometric and temperature adjustments), background correction, experimental SUIT (substrate-uncoupler-inhibitor-titration) protocols and data analysis.

With the O2k core technology, developed exclusively by Oroboros for high-resolution measurement of oxygen flux, the NextGen-O2k was developed.

O2k

High-resolution respirometry (HRR) and unique combinations of the O2k with

Fluorometry: the O2k-FluoRespirometer
Potentiometry: the O2k-pH ISE-Module and O2k-TPP+ ISE-Module
Amperometric measurement of nitric oxide: the O2k-NO Amp-Module

NextGen-O2k

Extend high-resolution respirometry (HRR) to the all-in-one device

The NextGen-O2k is a MultiSensor all-in-one instrument for in-depth studies of (1) the mitochondrial role in human diseases, and (2) mitochondria and chloroplasts ability in regulating algal growth and metabolite production rates.

(+1) Detection of the redox state of the Q-junction: MiPNet24.12 NextGen-O2k: Q-Module

(+2) Controlled wavelengths and light intensity: PhotoBiology-Module

Save on experimental time, reagents and samples
Minimize errors and ensure reproducible and accurate results
Highest oxygen sensitivity (5 nM detection limit)
Exceptional resolution (wide oxygen concentration range, between 0 - 1,000 μM)
Robustness (>10 years lifespan)

Objectives

The overall goal of this Phase 2 project is to complete the technical and scientific development of the new NextGen-O2k and prepare it for market launch.

WP1

» Technical developments

WP2

» Scientific developments

WP3

WP4

WP5

WP6

Coordinator

Oroboros Instruments GmbH

Network

Partners

Our partners WGT-Elektronik and HTech have been essential in the development of the NextGen-O2k.

Early testers

The influence of KOLs and Early testers/adopters will be used to build awareness of NextGen-O2k among researchers, academic and industrial partners. We have confirmed relevant mitochondrial and algal biotechnology researchers who want to collaborate as early testers:

Biomedical research	Algal research
Prof. Massimo Zeviani ^a,b, Dr. Carlo Viscomi ^c, Prof. Mervyn Singer ^a ^a Medical Research Council - Mitochondrial Biology Unit, University of Cambridge, UK ^b Department of Neurosciences, University of Padova, IT ^c Department of Biomedical Sciences, University of Padova, IT	Dr. Tomas Morosinotto Department of Biology, University of Padova, IT
Prof. Pablo Garcia-Roves Department of Physiological Sciences, Faculty of Medicine and Health Sciences, University of Barcelona and Bellvitge Biomedical Research Institute (IDIBELL), ES	Prof. Yagut Allahverdiyeva-Rinne Faculty of Technology, University of Turku, FI
Dr. Anthony Molina Division of Geriatrics and Gerontology, Department of Medicine, University of San Diego, US	Prof. Claire Remacle Genetics and Physiology of Microalgae, InBios/Phytosystems Research Unit, University of Liege, BE
Dr. Marten Szibor Clinic for Heart and Thorax Surgery, Jena University Hospital, DE
Prof. Howard Jacobs Institute of Biotechnology, University of Helsinki, FI
Dr. Alexander Karabatsiakis Institute of Psychology, Department of Clinical Psychology, University of Innsbruck, AT

NextGen-O2k events

	When	Where
NextGen-O2k Launch event 2021 Innsbruck AT	2021-10-13	Innsbruck AT, 2021 Oct 13-19. NextGen-O2k Launch event, NextGen-O2k SME2.
NextGen-O2k Event Photobiology: Algal bioenergetics Innsbruck AT	2021-07-15	Innsbruck AT, 2021 Jul 15-16. NextGen-O2k Event Photobiology: Algal bioenergetics.
World summit on Diabetes 2021 Virtual	2021-06-16	Vitual, 2021 Jun 16-17. World Summit on Diabetes: New technologies and practical approaches: Diabetes and Endocrine disorders
4th edition Metabolism & Cancer 2021 Virtual	2021-05-27	Virtual, 2021 May 27-29. 4th edition Metabolism & Cancer.
ISAP 2021 Virtual	2021-05-14	Virtual, 2021 May 14-Aug 13. 7th Conference of the International Society for Applied Phycology - ISAP2021.
Career Event - FH Campus 2021 Wien AT	2021-01-26	Virtual, 2021 Jan 26. Career Event of the FH Campus Wien.
Virtual OroDM02	2021	Virtual Oroboros Distributor Meeting, OroDM02
AlgaEurope 2020 Virtual Event	2020-12-01	Rome, IT, 2020 Dec 1-5. AlgaEurope 2020.
Long Night of Research 2020 Virtual Event	2020-10-09	Virtual Event, 2020 Oct 09. Oroboros at Long Night of Research, CCB. The diagnostic bioenergetic report – a milestone on the way to mitochondrial fitness and physical well-being.
36th Congress Czech Nutrition Society 2020 Hradec Kralove CZ	2020-09-10	Hradec Kralove, CZ, 2020 Sep 10-12 . 36th annual international congress of Czech Nutrition Society
MitoEAGLE WG2 Innsbruck AT	2020-03-09	Innsbruck AT, 2020 Mar 9-13. WG 2 on human skeletal muscle fibers retreat - COST Action MitoEAGLE.
MitoEAGLE WG4 Innsbruck AT	2020-02-26	Innsbruck AT, 2020 Feb 26-28. WG 4 blood cell workshop and retreat - COST Action MitoEAGLE.
MiPNet25.10 Distributor engagement Tokyo JP	2020-02-04	Tokyo,JP, 2020 Feb 4-7. On-site O2k-Workshop on High-Resolution Respirometry and partner distribution engagement.
MiPNet25.06 IOC145 Innsbruck AT	2020-01-27	Innsbruck AT, 2020 Jan 27-29. O2k-Coaching Days - Basic IOC145.
MiPNet25.03 IOC144 Innsbruck AT	2020-01-20	Innsbruck AT, 2020 Jan 20-22. O2k-Coaching Days - Basic IOC144.
1st Myocardial Function Symposium 2020 Graz AT	2020-01-10	Graz, AT, 2020 Jan 10-11. 1st Myocardial Function Symposium: “Targets in cardiometabolic disease”.
NextGen-O2k SME2 coaching Orily Pratt part 2	2019-12-03	Innsbruck AT, 2019 Dec 03-04 NextGen-O2k SME2 coaching Orily Pratt (part 2), NextGen-O2k SME2.
MiPNet24.15 IOC143 Haryana IN	2019-11-15	Haryana IN, 2019 Nov 15. Oroboros O2k-Workshop on high-resolution respirometry: Assessment of mitochondrial function in health and disease IOC143.
Neurocon 2019 Haryana IN	2019-11-15	Haryana, IN. 2019 Nov 15-18, Neurocon 2019.
MiP2019/MitoEAGLE Belgrade RS	2019-10-13	Belgrade RS, 13-16 Oct 2019. 14^th Conference on Mitochondrial Physiology: Mitochondrial function: changes during life cycle and in noncommunicable diseases - COST MitoEAGLE perspectives and MitoEAGLE WG and MC Meeting.
MiPNet24.13 IOC142 Fukuoka JP	2019-10-03	ASMRM & J-mit 2019, Fukuoka JP, 2019 Oct 03 Oroboros Lunch Seminar at ASMRM & J-mit 2019 Fukuoka JP - Explore the Mito-World! (IOC142).
ASMRM & J-mit 2019 Fukuoka JP	2019-10-03	Fukuoka, JP, 2019 Oct 03-05. Joint ASMRM and J-mit Conference
MiPNet24.02 IOC141 Schroecken AT	2019-09-23	Schroecken AT, 2019 Sep 23-28 Oroboros O2k-Workshop on high-resolution respirometry (HRR), IOC141.
NextGen-O2k SME2 coaching Samia Kappe part 1	2019-09-12	Innsbruck AT, 2019 Sep 10-11 NextGen-O2k SME2 coaching Samia Kappe (part 1), NextGen-O2k SME2.
FEBS Workshop Ageing 2019 Innsbruck AT	2019-09-09	Innsbruck, AT. 2019 Sep 09-12, FEBS Workshop “Ageing and Regeneration”.
Mitochondrial Physiology ‐ from Organelle to Organism 2019 Copenhagen DK	2019-08-19	Copenhagen DK , 2019 Aug 19-23. Mitochondrial Physiology ‐ from Organelle to Organism
NextGen-O2k SME2 coaching Orily Pratt part 1	2019-07-22	Innsbruck AT, 2019 Jul 22-23 NextGen-O2k SME2 coaching Orily Pratt (part 1), NextGen-O2k SME2.
MiPschool Coimbra 2019	2019-07-08	Coimbra, PT. 2019 Jul 08-11, 12^th MiP/MitoEAGLE Training School 2019. Mitochondrial respiratory physiology: Challenges on data sharing, reproducibility, and interpretation.
OroDM01 Innsbruck AT	2019-07-01	Innsbruck AT, 2019 Jul 01-03 Oroboros Distributor Meeting, OroDM01.
MiPNet24.04 IOC140 Amsterdam NL	2019-06-23	Amsterdam NL, 2019 Jun 23 Pre-conference Oroboros O2k-Workshop on high-resolution respirometry (HRR), IOC140.
MiPNet24.01 IOC139 Schroecken AT	2019-06-17	Schroecken AT, 2019 June 17-22 Oroboros O2k-Workshop on high-resolution respirometry (HRR), IOC139.

NextGen-O2k dissemination

NextGen-O2k press releases

Depression als Art 'Burnout' in den Zellen - Tiroler Tageszeitung online, Sep 2020, 2020-09-24
Rekord bei Patenten, Tirol auf Rang sechs - Tiroler Tageszeitung, July 2020, 2020-07-03
Oroboros Instrument revolutioniert Zellbiologische Wissenschaft - www.wirtschaftszeit.at; 2020-03-30
Beste Innovationen aus Tirol] www.economyaustria.at, 2019-12-16
Clusterawards 2019: Ausgezeichnete Innovationen aus Tirol science.apa.at, 2019-12-13
Tiroler Unternehmen für Innovationen mit Clusterawards 2019 ausgezeichnet - www.wirtschaftszeit.at; 2019-12-16
Clusterawards 2019: Ausgezeichnete Innovationen aus Tirol www.meinbezirk.at, 2019-12-13
Tiroler Forschungsunternehmen startet durch - Tiroler Landeszeitung, September 2019, S30-31, 2019-09-23
Innovationsförderung Tirol - Oroboros erhält Zuschuss in Millionenhöhe, Der Report 2019, Sonderheft der Wirtschaftsnachrichten, S12-13, 2019-09-18
Dem Leben beim Atmen zusehen Der Standard, Beilage Forschung spezial, S 13, 2019-08-07.
Turbo für Mitochondrienforschung und Oroboros Instruments, Standortagentur Tirol online, 2019-07-29
Tiroler Unternehmen Oroboros erhält 1.7 Millionen Euro EU-Förderung, tt.com, 2019-07-29
Turbo für Mitochondrienforschung udn Oroboros Instruments science.apa.at, online, 2019-07-29
1.7 Millionen Euro Förderung für Oroboros Instruments www.meinbezirk.at online, 2019-07-29
1,7 Millionen Euro EU-Flörderung für Tiroler KMU www.derbrutkasten.com online, 2019-07-29
Tiroler Erfindung macht in Europa Furore theworldnews.net online, 2019-07-29
Tiroler Erfindung macht in Europa Furore www.krone.at online, 2019-07-29
EUR 1,7 Mio Förderung für Tiroler KMU Oroboros Tiroler Tageszeitung, S 16, 2019-07-30
Tiroler Millionen-Erfindung Tiroler Krone, S 22, 2019-07-30

Gentle Science

In addition to O2k-Workshops and on-site training, Oroboros will disseminate the not-for-profit initiatives on Mitochondrial Physiology and Bioenergetics Science Camps, specialized Mitochondrial Physiology Conferences, and Preprint publications:

» Oroboros is a key player in support of the COST Action MitoEAGLE

» MitoGlobal - the world-wide platform for societies and organizations supporting mitochondrial research and medicine

» MitoEAGLE with cooperation by Oroboros launched MitoFit Preprints - the preprint server for mitochondrial physiology and bioenergetics

» A vision on preprints

» Mitochondrial respiratory states and rates : >530 coauthors join forces towards harmonization on nomenclature

» Gentle Science - recognizes the responsibility of the scientific community - for the quality of science, the quality of life in science, and its mission

MitoEAGLE Q-workshop

Electron supply to the Q-junction: assessment of mitochondrial respiration, H2O2 flux and the redox state of the Q-pool. COST Action MitoEAGLE, Innsbruck, Nov 2018.

Support

Supported by project NextGen-O2k which has received funding from the European Union’s Horizon 2020 research and innovation programme under grant agreement No. 859770.

MitoPedia: NextGen-O2k

» NextGen-O2k«

Term	Abbreviation	Description
Hydrogen peroxide	H₂O₂	Hydrogen peroxide, H₂O₂ or dihydrogen dioxide, is one of several reactive oxygen intermediates generally referred to as reactive oxygen species (ROS). It is formed in various enzyme-catalyzed reactions (e.g., superoxide dismutase) with the potential to damage cellular molecules and structures. H₂O₂ is dismutated by catalase to water and oxygen. H₂O₂ is produced as a signaling molecule in aerobic metabolism and passes membranes more easily compared to other ROS.
Light-enhanced dark respiration	LEDR	Light-enhanced dark respiration LEDR is a sharp (negative) maximum of dark respiration in plants in response to illumination, measured immediately after switching off the light. LEDR is supported by respiratory substrates produced during photosynthesis and closely reflects light-enhanced photorespiration (Xue et al 1996). Based on this assumption, the the total photosynthetic oxygen flux TP is calculated as the sum of the measured net photosynthetic oxygen flux NP plus the absolute value of LEDR.
Mitochondrial membrane potential	mtMP, Δψ [V]	The mitochondrial membrane potential, mtMP, is the electric part of the protonmotive force, Δp_H+. Δψ = Δp_H+ - Δµ_H+ / F mtMP or Δψ is the potential difference across the inner mitochondrial (mt) membrane, expressed in the electric unit of volt [V]. Electric force of the mitochondrial membrane potential is the electric energy change per ‘motive’ electron or per electron moved across the transmembrane potential difference, with the number of ‘motive’ electrons expressed in the unit coulomb [C].
NextGen-O2k
NextGen-O2k Instrument		NextGen-O2k Instrument
NextGen-O2k Technical developments
Oxygen kinetics		Oxygen kinetics describes the dependence of respiration of isolated mitochondria or cells on oxygen partial pressure. Frequently, a strictly hyperbolic kinetics is observed, with two parameters, the oxygen pressure at half-maximum flux, p₅₀, and maximum flux, Jmax. The p₅₀ is in the range of 0.2 to 0.8 kPa for cytochrome c oxidase, isolated mitochondria and small cells, strongly dependent on J_max and coupling state.
PB-Module	PB-Module	The PB-Module has been developed for conducting measurements of PhotoBiology, including photosynthesis. It consists of the PB-Sensor and electronic components which are an integral part of the NextGen-O2k. Measurements are recorded and evaluated with the DatLab 8 software.
PB-Sensor		The PB-Sensor has been designed as a part of the PB-Module to provide with an external source of light, in the range of the blue (451 nm) or red (634 nm) wavelengths. This enables experiments for evaluating the production/consumption of O₂ in the presence of light. The PB-Sensor consists on one LED and one photodiode mounted on the sensor tip behind a plastic cover. The LED emits with blue or red light on the sample and its intensity can be regulated from 0 to 5000 µEinsteinsm^-2s^-1. The photodiode provides real-time measurement of the light intensity allowing for continuous adjustment to the desired value, which can also help compensate for the temperature rise due to the operation of the LED.
PhotoBiology	PB	PhotoBiology is the science of the effect of light on biological processes. This includes photosynthesis, photochemistry, photophysics, photomorphogenesis, vision, bioluminescence, circadian rhythms and photodynamic therapy. Phototoxicity results from non-ionizing radiation (i.e. ultraviolet, visible and infrared radiation). Non-ionizing radiation is any type of electromagnetic radiation that does not carry enough energy per quantum (photon energy below 10 eV) to completely remove an electron from an atom or molecule. When photons interact with molecules, the molecules can absorb the photon energy and become excited, reacting with surrounding molecules and stimulating "photochemical" and "photophysical" changes. Respiration may be affected by light during photosynthesis or in dark respiration, with the transient response of light-enhanced dark respiration.
Q-Module	Q-Module	The Q-Module, developed both for measuring the Q redox state and for cyclic voltammetry measurements, is an integral part of the NextGen-O2k and consists of the Q-Sensor, integrated electronic components in the O2k, and the DatLab software.
Q-Sensor		The Q-Sensor has been designed as a part of the Q-Module for measurements with cyclic voltammetry and voltammetry, allowing for analysis of the Q redox state. The Q-Stopper with the reference electrode is called Q-Sensor, which is plugged in the NextGen-O2k. A three-electrode system is used to detect the Q redox state. Two of the three electrodes (glassy carbon and platinum electrode) are built into the Q-Stopper, while the reference electrode is removable (Reference-Electrode\2.4 mm).