Difference between revisions of "Troubleshooting: fluorescence module"
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Revision as of 12:12, 19 March 2021
Troubleshooting: fluorescence module |
MitoPedia O2k and high-resolution respirometry:
O2k-Open Support
In the following page, the most common problems occurring with either the O2k-Fluo Smart-Module or O2k-Fluo LED2-Module are summarized. The problem can be related to (a) the fluorescence settings in DatLab, (b) the fluorescence sensors, (c) the Fluorescence-Control Unit (in the case of the O2k-Fluo LED2-Module), or (d) the O2k.
Zero fluorescence signal
- Problem: The raw fluorescence signal is 0 V with and without the fluorescence dye.
- Solution: To identify cause of the problem, please go through the following check-list:
- 1. Check that the Fluo-Sensors are connected to the O2k and inserted into the O2k-chamber in the right position. See: O2k-Fluo_Smart-Module#Connect_Fluorescence-Sensor_to_O2k.
- 2. Check that the Fluorescence-Control Unit is properly connected to the O2k of series D-G.
- 3. In DatLab check that:
- a) the illumination is switched off in the ´O2k control´window.
- b) the gain for Fluo-Sensor and the Fluo intensity are set to a number greater than 0 in the 'Amperometric,Amp' tab of the ´O2k control´ window.
- 4. If all the above-mentioned settings and set up are correct and the detected signal with the Fluo-Sensor is still 0, perform the following test to localize the problem:
- Add distilled water or respiration medium into the O2k-chamber (without sample), insert the black stopper and change the settings for fluorometric measurements: Gain=1000; Fluo intensity (Serie H or higher) or Polarisation voltage (Series D-G)=500.
- 3. In DatLab check that:
- 1. run: Fluo-Sensor A in chamber A and Fluo-Sensor B in chamber B (let it run for 5 min)
- 2. run: Fluo-Sensor A in chamber B and Fluo-Sensor B in chamber A (let it run for 5 min)
- If the problem is with the Fluo-Sensor, the fluorescence signal will be 0 V with the faulty Fluo-Sensor in both chambers. In this case, the Fluo-Sensor should be shipped to us for repairs.
- If the problem is with the O2k or the O2k-LED2 Fluo-Module, 0 V fluorescence signal can be observed just in one chamber with two different Fluo-Sensors. In this case, the O2k or the Fluorescence-Control Unit (Series D-G) should be shipped to us for repairs. In some cases, the defect of the Fluorescence-Control Unit (Series D-G) leads to 0 V in both chambers.
- Please contact our O2k-technical Support to analyze your problem before shipment. Be sure to always send us original DatLab files with the above-mentioned tests to evaluate Fluo-Sensor performance.
Noisy fluorescence signal
- Problem: The raw fluorescence signal is noisy to such an extent that the evaluation of the results is not possible.
- Solution: To localize the problem, perform the following test:
- Add distilled water or respiration medium into the O2k-chamber (without sample), insert the black stopper and change the settings for fluorometric measurements: Gain=1000; Fluo intensity (Serie H or higher) or Polarisation voltage (Series D-G)=500.
- 1. run: Fluo-Sensor A in chamber A and Fluo-Sensor B in chamber B (let it run for 5 min)
- 2. run: Fluo-Sensor A in chamber B and Fluo-Sensor B in chamber A (let it run for 5 min)
- If the problem is with the Fluo-Sensor, the fluorescence signal will be noisy with the faulty Fluo-Sensor in both chambers. In this case, the Fluo-Sensor should be shipped to us for repairs.
- If the problem is with O2k or the O2k-LED2 Fluo-Module, the fluorescence signal is noisy 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 for repairs. If the noise occurs in both chambers, the Fluo-Sensors and the O2k (and Fluorescence-Control Unit - series D-G) should be shipped to us.
- Please contact our O2k-technical Support to analyze your problem before shipment. Be sure to always send us original DatLab files with the above-mentioned tests to evaluate Fluo-Sensor performance.
- Example: Problem with O2k or O2k-LED2 Fluo-Module
- Provided by Hazirah Hassan, University Kebangsaan Malaysia, [email protected]
- After changing the Fluo-Sensors between the O2k-chambers, 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 in chamber B, which makes the experiment hard to interpret. Both the O2k and the LED2 Fluo-Module should be shipped to our electronic workshop.
Negative fluorescence signal
- Problem: The fluorescence signal of one Fluo-Sensor is negative.
- Solution: To localize the problem, perform the following test:
- Add distilled water or respiration medium (without sample) into the O2k-chamber, insert the black stopper, and change the settings for fluorometric measurements: Gain=1000; Fluo intensity (Series H or higher) or Polarisation voltage (Series D-G) = 500.
- 1. run: Fluo-Sensor A in chamber A and Fluo-Sensor B in chamber B (let it run for 5 min)
- 2. run: Fluo-Sensor A in chamber B and Fluo-Sensor B in chamber A (let it run for 5 min)
- The problem is related to one of the Fluo-Sensors, if you observe a negative signal in both chambers after swapping them. The negative signal might be related to a wet plug of the Fluo-Sensor. The solution is to dry it out with e.g. a hair dryer and let it run connected to the O2k (25 °C, illumination off, distilled water in the O2k-chamber, gain for Fluo-Sensor 1000, fluorescence intensity=500) for a couple of days in a dry and warm environment. If this treatment does not solve the problem and you get still negative fluorescence values, the Fluo-Sensor should be sent back to us for further investigation.
- Please contact our O2k-technical support to analyze your problem before shipment. Be sure to always send us original DatLab files with the above-mentioned tests to evaluate Fluo-Sensor performance.
- Example: Provided by Anne Laure Charles, FR_Strasbourg_Zoll J,anne laure charles <[email protected]
- The fluorescence signal was negative in both chambers after swapping the Fluo-Sensors, but the signal changed after swapping the sensors, which means that the problem is located to the Fluo-Sensors. In this case, both Fluo-Sensors should be shipped to the electronic workshop.
Fluorescence signal is constantly at 10 V
- Problem: The fluorescence signal is constantly at the 10 V upper limit when using the O2k-Fluo Smart-Module or O2k-Fluo LED2-Module.
- Solution: This is probably due to high light exposure to the photodiode. Check the following parameters:
- 1. The chamber illumination must be switched off. Check it in the O2k control window.
- 2. The photodiode filters must be inserted, otherwise the light emitted from the LED will not be filtered. Information on how to mount the filter cap- » here.
- 3. Black PEEK stoppers must be used.
- 4. The fluorescence intensity cannot be set too high. Check in the Amperometric,Amp tab of the ´O2k control´ window.
- If all the above-mentioned settings and set up are correct and the detected signal is still at 10 V (out of scale), the O2k (and Fluorescence-Control Unit - series D-G) or/and the Fluo-Sensors should be shipped to us.
- Please contact our O2k-technical Support to analyze your problem before shipment. Be sure to always send us original DatLab files with the above-mentioned tests to evaluate Fluo-Sensor performance.