Troubleshooting: fluorescence module: Difference between revisions
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== | == Zero fluorescence signal == | ||
Β | ::: '''Problem:''' The raw fluorescence signal is 0 V with and without the fluorescence dye. | ||
::: '''Problem:''' The raw fluorescence signal is 0 V | |||
:::'''Solution:''' How can you identify cause of the problem? | :::'''Solution:''' How can you identify cause of the problem? | ||
:::: Please go through the following check-list: | :::: Please go through the following check-list: | ||
Line 17: | Line 16: | ||
:::::: If the problem is with the Fluo-Sensor, the fluorescence signal will be abnormal with the faulty Fluo-Sensor in both chambers. In this case, the Fluo-Sensor should be shipped to us for repairs. Β | :::::: If the problem is with the Fluo-Sensor, the fluorescence signal will be abnormal with the faulty Fluo-Sensor in both chambers. In this case, the Fluo-Sensor should be shipped to us for repairs. Β | ||
:::::: If the O2k or the O2k-LED2 Fluo-Module are defective, 0 V or noisy 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 for repairs. Β | :::::: If the O2k or the O2k-LED2 Fluo-Module are defective, 0 V or noisy 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 for repairs. Β | ||
::::: Please contact our [https://www.oroboros.at/index.php/o2k-technical-support/ 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:''' How can you 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. | |||
::::::* 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 O2k or the O2k-LED2 Fluo-Module are defective, 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. | |||
::::: Please contact our [https://www.oroboros.at/index.php/o2k-technical-support/ 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. | ::::: Please contact our [https://www.oroboros.at/index.php/o2k-technical-support/ 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: Defective O2k or O2k-LED2 Fluo-Module''' | :::: '''Example: Defective O2k or O2k-LED2 Fluo-Module''' |
Revision as of 14:49, 18 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: How can you 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. In DatLab check that:
- a) the illumination is switched off in the Β΄O2k controlΒ΄window.
- b) the gain for Fluo-Sensor and the Fluo intensity are a number greater than 0 in the 'Amperometric,Amp' tab of the Β΄O2k controlΒ΄ window.
- 3. 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.
- 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 abnormal with the faulty Fluo-Sensor in both chambers. In this case, the Fluo-Sensor should be shipped to us for repairs.
- If the O2k or the O2k-LED2 Fluo-Module are defective, 0 V or noisy 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 for repairs.
- 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: How can you 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.
- 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 O2k or the O2k-LED2 Fluo-Module are defective, 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.
- 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: Defective 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. 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.
Negative fluorescence signal
- Problem: The fluorescence signal of one Fluo-Sensor is negative, while the other Fluo-Sensor has ~0 V.
- Example: Provided by Anne Laure Charles, FR_Strasbourg_Zoll J,anne laure charles <[email protected]
- Solution: 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 (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)
- 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 Β°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 and replacement. 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.
Fluorescence signal is constantly at 10 V
- Problem: The fluorescence signal is constantly at 10 V 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:
- 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. Information on how to mount the filter cap- Β» here.
- Black PEEK stoppers must be used.
- The fluorescence intensity cannot be set too high. Check in the Amperometric,Amp tab of the Β΄O2k controlΒ΄ 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 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.
Useful links
- Β» Amplex UltraRed
- Β» Safranin
- Β» TMRM
- Β» Rhodamine 123
- Β» Magnesium Green
- Β» Calcium Green
- Β» O2k-FluoRespirometer manual
- Β» O2k-Fluo LED2-Module
- Β» O2k-Demo experiments
- Β» Sole source information
- Β» Oroboros O2k