Bioanalyzer-Agilent Protein 80 Error

I've been using Protein 230 Kit for a while without any issue. Then I tried Protein 80 Kit, but I immediately got an error message. The instrument is about 10 years old but I replaced new cartridge. It's working so good with Protein 230. I did the Diagnosis but failed in the last three because I have no those special chip.

1|Instrument error occurred on port 1, Unusual high or low voltage or current was detected during the start phase of the on-Chip analysis.
Wells marked with (+) or (-) have been causing problems. The top left well equals sample 1 on the microfluidic chip:

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  • Hello MDAZ,

    Thank you for contacting the Agilent team.

    The unusual high low voltage error you are seeing is often a routine cleaning related issue and not related to instrument hardware per se. However, to be sure that instrument hardware is fine, full set of hardware diagnostics can be run using the test chip kit P/N G2938-68300. The unusual high low voltage error occurs when one or more of the electrodes are making poor connection with the liquid in the wells. For example, an empty well, or a well that has insufficient volume can trigger this error. Although Protein 230 assays have worked normally for you and the issue is seen when running Protein 80 assays, addressing the common causes of the issue is in order. Therefore, please refer to page 96 of the 2100 Bioanalyzer Maintenance and Troubleshooting Guide (M&T guide) for likely causes and steps to address this issue. Specifically, please address the following points: 

    1. Clean the pin-set (electrode cartridge). As first step, please thoroughly clean the pin-set using a baby toothbrush following the detailed protocol as described starting page 126 of the M&T guide. Then ensure that it is thoroughly rinsed to remove any traces of sample, RNAse ZAP from the pin-set if that was used for cleaning. Then make sure to dry the pin-set from front and back before putting it back on the instrument and then do the 'short circuit test' as described on step 14 of page 130 of the M&T guide. This test requires a new empty chip and not a test chip. For pin-set maintenance, you can also refer to ‘How to maintain the electrode cartridge video ‘.
    2. Clean the priming station. Priming station serves the purpose of pushing gel through the microfluidic channels so this should work optimally. This is also one of the main culprits in suboptimal migration issues. Therefore, please clean/change parts of the priming station if those did not get changed recently. The cleaning procedure is outlined in the M&T guide starting page 131 and below table shows the frequency of the recommended maintenance for the priming station:


    Particularly, the black adapter can be cleaned with a 27-gauge needle. Any gel buildup would easily come out with this. After the cleaning is done and the priming station is assembled, please perform seal test for the priming station prior to your next run. The purpose of seal test is to verify performance of the priming station. For priming station maintenance, you can also refer to ‘ How to Maintain the Priming Station video ‘.

           3.  Perform a run. After executing the above listed steps, please verify the maintenance by doing a run. Before the run:

    • Please make sure that the reagents are allowed to fully equilibrate at 25°C for 30 minutes.
    • Please prepare fresh gel dye mix using an unexpired kit
    • Please ensure that base plate and syringe clip settings are correct for Protein 80 assay (see below).


    Please let us know if there are any questions. Also, if further extensive troubleshooting is needed, please contact our support team for US and Canada at

    Thank you, Shweta

  • Hi Shweta,

    Thank you for your help. It worked after I cleaned everything. Then I was very disappointed with Protein 80 Kit. I have attached the results. I have two samples both with two peaks around 30KDa; and I need to calculate the ratio of these two peaks and I actually don't care much about the weight measurement. With Protein 230 Kit there was little separation for sample #1 and I can calculate an approximate rate, but with Protein 80 Kit only one peak was shown. The same thing happened with sample #2. The Protein 80 Kit should give a better separation than the Protein 230 Kit. But I got an opposite result. Do you have any suggestion? How do I tell Software to separate if the peak has overlapping with ladder?

  • Hi MDAZ, Thank you for the update that the high low voltage issue is resolved.

    Regarding issue related to size resolution using Protein 80 verses Protein 230 kit, may I request you to email corresponding raw data files ending with .xad to the Americas support team at . Default location is C:\Program Files (x86)\Agilent\2100 bioanalyzer\2100 expert\data.

    Unfortunately, there is no way to change settings in the software to separate peak. However, by looking at raw data files we can determine how to best address the issue.

    Thank you, Shweta

  • Hi Shweta,

    Thank you. I followed your advice and sent the raw data. Hopefully we can figure it out.

  • Thank you for the update. I checked, our team has received your query in order and your are being assisted. Thanks again. 

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