I am not convinced that existing markers are useful to address such a question and it depends on what level of hypoxia you are interested in examining. Most endogenous markers depend on activation of HIF1 and of course cellular location may be affected by transport of the molecule in the cell. Most endogenous markers such as pimonidazole are metabolised to a product that binds in the cell but this usually occurs only at significantly lower oxygen levels than are required to upregulate, again products may diffuse before binding. Suggest that you think about another approach. See for example O'Loughlin et al Rad Res 1980, 84(3) 477-95.

HIF is translocated to the nucleus upon activation in a hypoxic environment so if you try nuclear extracts on a western blot you might get a more specific signal of hypoxia.

Are you using a cell culture system?

I agree with Richard. You would need targeted indicators, and as far as I am aware there are none that would do the job. It also depends on what you mean by core. The lowest PO2 will be within the mitochondrial matrix, then the perimitochondrial region. There have been a lot of studies in various tissues where the mitochondrial PO2 has been estimated from mitochondrial function, membrane potential and the like, but these are highly indirect and open to challenge.

I agree with Jeremy. There is an absence for a specific marker to indicate cell ischemia.

Richard is right. Molecular "markers" as HIF, RISK, p-Akt etc do not mark "hypoxia" but rather the cell response to hypoxia. Suppose you have a human red cell: if you put it in an anoxic environment, does it over-express HIF and Co? The answer is clearly no, but the cell is nevertheless "hypoxic". Moreover, various cell lines respond to different extents depending on many factors as their own metabolic activity, their own oxygen extraction and many others, but the degree of hypoxia is the same. Likewise, I expect that the various intracellular compartments may follow the same rules. I believe that the question should be reformulated. The response to ischemia may be somewhat different from that to hypoxia, but I suspect that the basic rules are more or less the same.

Thank you all my dear colleagues for you invaluable Tips. I guess i must have to clarify more. Since we can not measure the intracellular hypoxia levels in different zones , we have no option but to measure the response to hypoxia . In this case i'm trying to find out wether oxygen can penetrate deep into the cell core ( center of the cell ) or not . this is an study about the cell size and possible hypoxic core inside the cell . So I'm waiting for your new suggestions, like a way to measure PO2 measurement inside the cells or something like that .... which i'm sure would be helpful.

Hadi, the only thing I could suggest is to use imaging of mitochondria function (e.g. potential or cytochrome redox status) in different regions of the cell, but this would be open to all sorts of problems and criticisms, as the potential dyes are by no means perfect and detecting cytochrome redox not easy, mitoPD varies a lot, and peripheral mitochondria may be different anyhow. The only thing you could do is to show the effects of going from hyperoxia to normoxia to hypoxia, and see if there are differential changes between the core and periphery. I don't think I would want to do this...

I suggest that you look into the possible use of oxygen sensitive phosphorescent probes. Try looking up the papers of David Wilson and sergei Vinogradov

look at B7Hi and tell me what you found

You may think about the level of O2 reduction to H2O at the level of electron transport chain in mitochondria. I think this can be measured through measuring the activity of the enzymes of that chain. at the surface of the cell, meaning cyctosol, give a glimpse if oxygenated myoglobin can assist for your purpose