I mean that the best way is ask about it clinician physiologist or peoples of laboratory of Haematology. I konow that under surgery to do non-stop monitoring of O2/CO2/OxyHb

Good luck!

There is near IR detection of oxyHb. That's your best chance. Other advanced techniques include Seahorse, which is expensive. 

Thanks for the reply. i wanted to know if there are any biochemical methods other than IR or hemox analyser.

I have emailed you a paper, plese check it might be useful

Measurement of oxygen saturation is easy with a blood gas analyzer. Oxygen content may be calculated by multiplication of saturation (as fraction) with Huefner´s number (1.36 ml O2/g Hb). Another possibility is measurement of oxygen content e. g. with a coulometric method ( Böning D et al. Respir. Physiol 34, 195-207 (1978).

calculate MCHC = Hb(in g/dl)/Hct,expressed in g/dl;

each gram of hemoglobin holds 1.34ml of oxygen

This is the method to calculate the bound form of oxygen in a given volume of packed red cell/red cell concentrate.

It depends on how much resources you have, and what you are actually aiming to do, in regards to your study protocol? You mentioned in your question RBC's in vitro. Are they freshly drawn RBC's? What species do they come from? Is the sample centrifuged? What medium are they suspended in?  Each method has its pro's and con's. Using the blood gas analyzer would probably be the best method, provided you have a blood gas analyzer handy. Most Emergency Departments have rapid analyzers around, which give an ABG in 5 minutes. If you want to measure real-time Hemoglobin changes, which allows you to measure the delta change in oxy/deoxy/total (µmol/L)and Tissue 02 saturation %, Maybe it would help if you gave a bit of background about what it is you want to achieve. Again, depends on the cost and set up. 

Thanks everyone for the response. I need to measure oxygen holding capacity of human RBCs, under completely oxygenated or under 5% CO2 condition, invitro. I can bubble pure oxygen to saturate RBCs with oxygen(oxyhemoglobin). then determine what is the maximum oxygen holding capacity of RBCs. Alternately, i can mimic tissue scenario, by bubbling 5% CO2 (deoxygenated condition. I can mix the 2 conditioned RBCs in different ratios to get a calibration curve.

However, I have constrains with the measurement tools.

One of the method is IR based absorbtion, or oxymeter, which i do not have in my lab. 

Are there any possibilities of using colorimetric method for determining oxygen saturation in blood (purified human RBCs).

Thanks Dieter Böning, for the paper. It looks you have done it. But its OK for me even with relative quantification.

I have heard that even i can use absorbtion from 500nm-600nm, where oxygenated hemoglobin has a distinct absorption maxima. Your views.

Invivo i can do it very easily with oxymeter. or any reflectance based measurements. But in isolated RBCs, how can i perform the experiment. 

u can measure the level 2,3-Bisphosphoglyceric acid  (2,3 DPG) level in isolated RBC. haemoglobin releases oxygen controlled by 2,3 DPG.

I need very specific measurement. I need to measure oxygen content in RBC

sir, it is biochemical method. other than u can do with blood gas analyzer...

I do not have blood gas analyser