Difficult cytokines

Alternatives Antibodies Cell type Difficult cytokines Fluorochromes Other species Protocol References Technique

 

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IL-1 The detection of interleukin-1 beta (IL-1) produced by monocytes asks for a different approach than usual because the production of IL-1 is post-transcriptionaly regulated. The pro-IL-1 is already present in the cytoplasm and clipped by ICE to 'produce' the bioactive IL-1. There will always be a background staining in the unstimulated sample and there is no point using a transport inhibitor. The staining will be diffuse throughout the cell and is difficult to distinguish from an aspecific binding of the antibody.

IL-10 One of the cytokines that is looked for in T lymphocyte immunology at the moment is IL-10. This cytokine is playing an important role in the regulation of the immune response. And it is very difficult to detect it intracellularly. The reports about this difficulty in detection are very heterogeneous. Some state it is not that difficult, some state it is impossible. In our experience it seem to be the (T) cells that are the most important factor in being able to detect IL-10.

Figure.1. Simultaneous detection of intracellular detection of IL-4, IL-10 and IFNy in human CD4+ T lymphocytes after 6 hrs PMA/ionomycin restimulation. In panel A. cells have had an initial neutral MLR stimulation (0.8% IL-10) and in panel B. the cells have had the same stimulation in the presence of a regulatory CD4+ T lymphocyte inducing factor (3.8% IL-10). In panel B. it is also clear that the majority of the IL-10 producing T lymphocytes also produces IL-4 (2.7%). From these cells app. 0.3% also produces IFNy.
 

IL-12 Also the detection of IL-12 is a special point of attention. We go into this under dendritic cells

IL-18 The same results can be obtained by the search for the cytokine IL-18. We regret the fact that there is only one company in the world that has the right to sell anti-human IL-18. And they do it at a price that is unrealistic. Many of us make the choice not to look into this cytokine.

TGF for instance is a molecule that is still difficult to detect. It is said (and published by M.L. Garba and J.A. Frelinger of UNC Chapel Hill, USA) that the clone TB21 is able to detect TGF. We did test this antibody and had no luck in detecting TGF in PBMC. We have found no other references than Garba et al. or other antibodies that show intracellular staining for TGF.

 

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Page was last updated: 05-03-2005