Dear collegues, I got the impression that in this ongoing discussion two different phenomena got mixed up. The one phenomenon that Maryalice described is the unspecific binding due to substances in the blood sample that cause unspecific binding of some but not all antibodies in your panel. It disappears by washing before staining or by adding access amounts of normal mouse serum. We see it in about one in a hundred cases esp. in the BD CD3-FITC/CD19-PE and CD3-FITC/CD16&56 combination. You see only two clusters in the two colour dot plot (see the quoted Cytometry paper). The other phenomenon of CD4+8+ has first been described to my knowledge by Diether Recktenwald in the application part of the first edition of the FACScan manual (it was removed later on). There he used it as an example for a three colour application where these double positive cells are propidium (or 7-AAD, I donīt remember exactly) positive. I have seen a few patients, where in all cases the normal mutual exclusive CD4 and CD8 are present and the third cluster of double positives in addition (in our hands in the BD CD4-FITC/CD8-PE test tube). The percentage of CD4+8+ was very stable in the two cases where we were able to get consecutive tests over a longer period of time. Interestingly, the first case was also a women from the lab where I did my PhD who was on thyroid hormones for the same reason. Others were outside patients where we could not get more clinical information. Taken all information together, apoptotic escapees form the thymus would be an attractive explanation. As these self reactive cells shouldnīt leave the special thymic environment, the attribution to autoimmune diseases is not a bad idea. I stop it here and apologize for my long comment I sent before. Thomas Nebe Klinikum Mannheim
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