Troubleshooting: fluorescence module |
MitoPedia O2k and high-resolution respirometry:
O2k-Open Support
Defective Fluo-Sensor or O2k or O2k-LED2 Fluo-Module
- If the raw fluorescence signal is 0 V with and without fluorescence dyes, it might be possible that your Fluo-Sensor is defective. How can you identify the problem?
- Please go through the following check-list:
- 1. Check that the illumination is switched off in the ´O2k control´window.
- If the raw fluorescence signal is 0 V with and without fluorescence dyes, it might be possible that your Fluo-Sensor is defective. How can you identify the problem?
- 2. Check the settings for the Amperometric,Ampchannel in the ´O2k control´ window, in the Amperometric,Amp tab. Gain for Fluo-Sensor and the Fluo intensity cannot be 0.
- 3. The Fluo-Sensors are connected to the O2k and inserted into the O2k-Chamber in the right position (see above in the 1.3 Connect Smart Fluo-Sensor to O2k section).
- 4. If all the above-mentioned steps are checked and the detected signal with your Fluo-Sensor is 0 (negative or very high) in order to localise the problem, perform the following test. Add distilled water or respiration medium into the O2k-Chamber, insert the black stopper and change the settings for fluorometric measurements (see above):
- 1. run: Fluo-Sensor A in chamber A and Fluo-Sensor B in chamber B
- 2. run: Fluo-Sensor A in chamber B and Fluo-Sensor B in chamber A
- If your Fluo-Sensor A is defective, the fluorescence signal should be 0 V with Fluo-Sensor A both in chamber A and B. In this case, the Fluo-Sensor should be shipped to us in order to repair it.
- If the O2k or the O2k-LED2 Fluo-Module are defective, 0 V or negative fluorescence signal can be observed just in one chamber with two different Fluo-Sensors (see example below). In this case, the O2k or the Fluorescence- Control Unit (Series D-G) should be shipped to us in order to repair it.
- Please contact our customer support to analyse your problem before shippment. Please always send us original DatLab files with the above-mentioned tests to evaluate the defective Fluo-Sensor performance.
- Problem: Defective O2k or O2k-LED2 Fluo-Module
- Provided by Hazirah Hassan, University Kebangsaan Malaysia, [email protected]
- Analysing these tests after changing the Fluo-Sensors between the O2k-Chambers, it is obvious that the problem is related to the O2k or O2k-LED2 Fluo-Module, since the fluorescence signal is negative and unstable in chamber B with both Fluo-Sensors. In the third figure the fluorescence signal is very unstable and it makes measurement unpossible with biological sample. Both the O2k and the LED2 Fluo-Module should be shipped to our electronic workshop.
- 4. If all the above-mentioned steps are checked and the detected signal with your Fluo-Sensor is 0 (negative or very high) in order to localise the problem, perform the following test. Add distilled water or respiration medium into the O2k-Chamber, insert the black stopper and change the settings for fluorometric measurements (see above):
The raw fluorescence signal [V] appears constantly at 10 V
- If you see a constant raw signal at 10 V when using the O2k-Fluo Smart-Module or O2k-Fluo LED2-Module, this is probably due to high light exposure to the photodiode. Check the following parameters:
- The chamber illumination must be switched off: Check it in the O2k control window.
- The photodiode filters must be inserted, otherwise the light emitted from the LED will not be filtered. Click here more information on how to mount the filter cap.
- Ensure that the Smart Fluo-Sensor is correctly inserted at the O2k frame of the chamber window.
- Use black PEEK stoppers.
- If you see a constant raw signal at 10 V when using the O2k-Fluo Smart-Module or O2k-Fluo LED2-Module, this is probably due to high light exposure to the photodiode. Check the following parameters:
Negative fluorescence signal
- Problem: The fluorescence signal of the Fluo-Sensor in chamber B is negative, while the other Fluo-Sensor has ~0 V. The same result occurs when the O2k-LED2 Fluo-Module is disconnected from the the O2k/switched off.
- Provided by Anne Laure Charles, FR_Strasbourg_Zoll J,anne laure charles <[email protected]
- Answer: Connacting our electronic workshop, the problem might be related to a wet plug of the Fluo-Sensors. The solution is to dry it out somehow e.g. hair dryer and let it run connected to the O2k for a couple of days in dry and warm environment. If these treatments cannot solve the problem and you get still negative fluorescence values, Fluo-Sensor should be sent back to us and our electronic partner will look at it and replace it if needed.