O2k-Open Support alert

»Oroboros Ecosystem«

O2k-Open Support alert - 2021

Currently Answered Questions from the »Oroboros Ecosystem Agenda«

Oct 2021

Oct-28 Th

New excel templates for H₂O₂ flux analysis with the Amplex UltraRed assay

- » H₂O₂ flux Excel templates« and »MiPNet24.10 H₂O₂ flux analysis«
- communicated by »Komlodi Timea« and »Cardoso Luiza«

Sep 2021

Sep-29 We

New excel templates and MiPNet for ATP production analysis with the Magnesium Green assay

- »MgG Excel templates« and »MiPNet26.10 MgG data analysis«
- communicated by »Cardoso Luiza« and »Komlodi Timea«

Sep-22 We

New excel templates for mitochondrial membrane potential analysis using fluorescence dyes

- »mt-membrane potential Excel templates« and »MiPNet24.09 General Template for Mt-membrane Potential Analysis«
- communicated by »Komlodi Timea« and »Cardoso Luiza«

Sep-09 Th

Can I compare Amplex UltraRed (AmR) experiments in which the H₂O₂ sensitivity is different?

The H₂O₂ sensitivity is calculated to calibrate the raw fluorescence signal to H₂O₂ equivalent concentration. Using the same gain and fluorescence intensity it is possible to compare experiments with different sensitivities.
- »AmR assay H₂O₂ sensitivity«
- communicated by »Cardoso Luiza« and »Komlodi Timea«

Aug 2021

Aug-26 Th

Is it normal to see a pink coloring in the medium at the end of an experiment with Amplex UltraRed (AmR)?

The pink coloring is due to the accumulation of UltroxRed, generated from Amplex UltraRed reacting with H₂O₂.
- »AmR assay and color of the medium«
- communicated by »Cardoso Luiza« and »Komlodi Timea«

Aug-19 Th

Which chemicals can I use in an experiment with Amplex UltraRed (AmR)?

Certain substances are incompatible due to their effect on the AmR assay sensitivity. Testing this effect before use is recommended.
- »Chemical compatibility with AmR«
- communicated by »Cardoso Luiza« and »Cecatto Cristiane«

Aug-12 Th

Do you know how to minimize the dilution of the AmR assay components upon sample addition?

Learn more about our recommendations:
- » AmR assay dilution «
- communicated by »Komlodi Timea« and »Cecatto Cristiane«

Aug-05 Th

What is the advantage of using flux control ratios (FCRs)?

FCRs provide information on coupling and pathway control of mitochondrial respiration independent of (i) mt-content in cells or tissues, (ii) purification of isolated mitochondrial preparations, and (iii) determination of tissue mass or mt-markers.
- »Flux control ratio«
- communicated by »Cardoso Luiza« and »Cecatto Cristiane«

Jul 2021

Jul-29 Th

Do you know how to export a user-specific DL-protocol (*.DLPU)?

- »Export_DL-Protocol_User_(*.DLPU)« and »O2k-Videosupport:Export DL-Protocol User«
- communicated by »Schmitt Sabine«, »Cecatto Cristiane«, and »Komlodi Timea«

Jul-28 We

Need to tailor a SUIT protocol to answer your research question?

See how to modify existing DL-protocols.
- »Enable_DL-Protocol_editing«
- communicated by »Schmitt Sabine«, »Cecatto Cristiane«, and »Komlodi Timea«

Jul-21 We

Not sure which SUIT protocol to apply?

The SUITbrowser is the perfect tool to find the best SUIT protocol for your research question.
- » SUITbrowser« and »O2k-Videosupport: How to find a DL-Protocol«

- communicated by »Schmitt Sabine« and »Komlodi Timea«

Jun 2021

Jun-28 Mo

Do I need to calibrate the fluorescence signal with safranin?

It is possible to run the experiment without calibration, presenting only the fluorescence intensity plot. However, a simple two-point or multiple-point calibration for safranin concentration is possible.
- »Safranin calibration«

Jun-16 We

Simultaneous fluctuations in O2 slope neg. and the signal of the Peltier power?

This can indicate an electrical disturbance and changing the electrical supply may resolve this problem.
- »Troubleshooting«
- communicated by »Cecatto Cristiane« and »Schmitt Sabine«

Jun-07 Mo

Do you observe high fluorescence signal even before adding the dye or sample?

Filters that have shifted from the correct position in the Fluo-Sensors might give rise to higher fluorescence signal than usual.
- »Troubleshooting: fluorescence module«
- communicated by »Komlodi Timea« and »Schmitt Sabine«

Jun-03 Th

How to ensure quality control when exchanging the 2-mL and the 0.5-mL chambers

Every time you exchange the chambers, a volume calibration and instrumental O2 background test should be performed.
- »MiPNet22.11 Section 5.4. O2k-chamber volume calibration« and »Oxygen flux - instrumental background«
- communicated by »Komlodi Timea« and »Cecatto Cristiane«

May 2021

May-25 Tu

Do you enter the correct instrumental background correction in DatLab for the experimental O2 concentrations?

After performing the instrumental background test at the appropriate O₂ range, load the parameters in DatLab to ensure accurate correction for external fluxes.
- »O₂ background correction in DatLab

- communicated by »Komlodi Timea« and »Cecatto Cristiane«

May-17 Mo

Planning to measure proton flux or mitochondrial membrane potential with ion-selective electrodes in the O2k?

Learn how to assemble the reference electrodes:
- »O2k-Videosupport: Assembly of the reference electrode«
- communicated by »Cardoso Luiza« and »Schmitt Sabine«

May-04 Tu

Do you know how to apply the air and zero calibration values in DatLab?

This important quality control step ensures accurate measurement of respiration.
For details on how to apply air and zero calibration in DatLab, see:
»Oxygen calibration - DatLab« and »Troubleshooting«
- communicated by »Komlodi Timea« and »Cecatto Cristiane«

Apr 2021

Apr-27 Tu

Why is the fluorescence signal constantly at 10 V or at 0 V?

Follow these steps to localize and solve the problem:
- »Fluorescence signal is constantly at 10 V« and »0 V fluorescence signal«

Apr-20 Tu

Noisy fluorescence signal?

Follow these steps to localize and solve the problem:
- »Noisy fluorescence signal«

Apr-13 Tu

Do you know what to do when the fluorescence signal starts at a negative value?

A negative signal may be caused by wet Fluo-Sensors. For further information:
- »Negative fluorescence signal«

Apr-06 Tu

A damaged OroboPOS-Holder may cause medium to leak from the O2k-Chamber.

See how you can check that the OroboPOS-Holders are intact.
- »O2k-OroboPOS-Holder«

Mar 2021

Mar-25 Th

O2k-checklist: Get started with an O2k-experiment

The O2k-checklist provides an overview of all essential steps for starting an O2k-experiment.
- » MiPNet20.04_O2k-checklist«
- » communicated by »Cecatto Cristiane« and »Schmitt Sabine«

Feb 2021

Feb-17 We

Do you know how to prepare permeabilized muscle fibers?

The application of high-resolution respirometry in permeabilized muscle fibers provides a sensitive diagnostic tool for the analysis of mitochondrial (dys)function in small biopsy samples.
- »O2k-Videosupport: Fiber permeabilization«
- »Doerrier 2018 Methods Mol Biol«
- communicated by »Doerrier Carolina« and »Tindle-Solomon Lisa«

Feb-08 Mo

Do you know how to properly insert the ISS-tip into the O2k-chamber?

With the stirrer bar still rotating, insert the ISS-tip to the bottom of the O2k-Chamber, so that it points away from you.
- »MiPNet19.03 O2k-cleaning and ISS: Section 2«
- communicated by »Doerrier Carolina«, »Cecatto Cristiane« and »Komlodi Timea«