Seahorse Xfe96 mito stress maximum OCR

Hi Rebecca,

Thank you for contacting Cell Analysis Technical Support.  We recommend maximum basal rates for 96 well platforms to be 160pmol/min. You may have potentially too many cells/well. If you could send your Wave file to Cellanalysis.support@agilent.com, we would be happy to take a look for you! How many cells did you seed per well and what type of cells are these?

Thank you!

Courtney Nadeau Watts

Technical Support Scientist

Cell Analysis Products

Hi Courtney,

Thanks for your quick response! I do believe we had too many cells per well. We are using rat primary cortical neurons at 80 and 100k cells/ well because in many cases a large portion of the neurons do not differentiate. We had previously done a test of cell density where the basal OCR looked fine at the same cell density but I think between different neuron preps we had different numbers of neurons that successfully differentiated. 

I am mostly wondering if there is any utility to the data as just a preliminary finding if the basal OCR is high or if the data is totally unusable in that case?

Thanks again,

Rebecca

Hi Courtney,

Thanks for your quick response! I do believe we had too many cells per well. We are using rat primary cortical neurons at 80 and 100k cells/ well because in many cases a large portion of the neurons do not differentiate. We had previously done a test of cell density where the basal OCR looked fine at the same cell density but I think between different neuron preps we had different numbers of neurons that successfully differentiated. 

I am mostly wondering if there is any utility to the data as just a preliminary finding if the basal OCR is high or if the data is totally unusable in that case?

Thanks again,

Rebecca

Hi Rebecca,

You are very welcome!  It is hard to say without seeing the data, unfortunately. We make the recommendation of 160pmol/min as the maximum because when it starts higher than that, once the FCCP is injected, the cells begin to work so hard that most/all of the O2 in the wells will be consumed, resulting in anoxic cells.  If you would like to send me the data file, I can better advise you!

Thank you,

Courtney