Sanne Hoeks

Chapter 6 84 FIGURE 2. Cytokine expression in various blood drawing tubes. Blood samples were obtained from healthy individuals using various blood collection tubes. After centrifugation cell free plasma (sodium heparin, citrate EDTA) and serum was measured using a multiplex immunoassay as described elsewhere . 15 Color profiles were generated using geometric mean values as previously described . 30 In general all different tubes types can be used for this kind of assay, though slight variations are observed between various blood collection tubes. For many clinical trials, experiments do not require the separation of blood components thus, most sample handling errors occur during the culturing of whole blood samples. To prevent the introduction of manipulation errors under these conditions a standardized culture system such as the TrueCulture© syringe can be used. 31 In this culture model, blood is collected directly from donors into a specially designed syringe tube. These tubes can be stored stably at -20 °C and can easily be thawed prior to use. The culture system creates a sealed environment in which freshly drawn blood can mix with a pre-prepared nutrient and stimulant solution. Samples can easily be incubated at 37 °C and once this incubation period is over, sedimented cells can be separated rapidly from the supernatant, preventing any further contact between the two environments. The syringes can be stored at -20 °C until further analysis, further minimizing analyte degradation. Data shows that cytokine levels measured using this system have a high degree of stability, with similar levels being measured in consecutive healthy donor blood draws. 31 Thus, in a clinical setting, this system facilitates stable and comparable culture conditions with minimal methodological error. Temperature and time delay of processing It is known that artifacts in cytokine measurements are affected by the duration of contact between serum or plasma and blood cells. 32 When plasma or serum cannot be isolated from whole blood or, if there is a delay in this separation procedure, it is important to understand cytokine dynamics under these conditions as changes here may greatly affect the experimental outcome and data analysis. For whole blood it has been shown that cytokine production can already occur within a 2-h period following blood draw. 33 When whole blood is stored at 4 °C, room temperature (RT) and 35 °C before separation into serum and plasma, some poignant changes in cytokine levels can be seen. 34 From a panel of inflammatory markers, keeping