Short intro: Our group is working on a novel dialysis device that uses electro-oxidation to remove metabolic waste products that are not easily bound by sorbent methods (mainly urea). In this way we hope to regenerate the dialysis fluid, making dialysis more portable.
In principle the unit works by passing blood by a regular dialysis membrane to filter out blood cells and molecules > 60 kd or so. The ultrafiltrate containing most of the toxins is then passed through an electro-oxidation unit that - for lack of a better way to put it - wreaks havoc on all molecular bonds. The oxidized or gasified degradation products are then filtered out using sorbents.
As you can imagine this quite a brute force approach and we want to see how toxic the oxidized and filtered plasma filtrate is at the end of the process (where ultimately it will be reintroduced in the patient circulation). We only have a broad idea of what kind of compounds are formed in the process and therefore don't know where to look for toxicity. The most likely mechanism would be oxygen radical generation. So, how would you approach this question?
We're currently looking at morphological blood abnormalities, such as hemolysis or the formation of abnormal RBC but haven't really seen much of it. The next step would be looking at lymphocyte viability by looking at WBC subsets on flow cytometry and measuring viability (and perhaps Annexin V staining).
Is the there a good way to look at ROS in WBC subsets? I was thinking to use CM-DCFDA as I've got lots of experience with it using cultured cells, but I'm not sure it'll work. I could also measure TBARS perhaps?
We should probably also look into lymphocyte activation (i.e. neutrophil degranulation, monocyte adhesion, interleukin production and...?)
So, has anyone done anything like this before? Any tips for assays (preferably with protocols)? What would you want to see if your were a regulator, journal reviewer or trial patient?
I suggest you to contact Prof. Joerg Vienken on this topic. He is a member here on Research Gate.
Thanks for the suggestions both of you. I'm getting fairly decent results with the phagoburst assay or with simpeler amplex red assays. Perhaps I'll contact Prof. Vienken for some additional ideas though.
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