Involvement of CD26 in Differentiation and Functions of Th1 and Th17 Subpopulations of T Lymphocytes

Figure 3

Separation of CD26^low and CD26^high lymphocytes by magnetic cell sorting (MACS). (a) Procedure of separation of CD26^low and CD26^high lymphocytes by MACS. After stimulation for three days, lymphocytes were labeled with the mouse anti-human CD26 mAb (anti-CD26 mAb₃₅₀ prepared in our own laboratory) for 1 h at 4°C. Following the two washing steps, magnetic MicroBeads labeled with anti-mouse IgG were added to the cells and incubated further for 15 min at 4°C. After a washing step, cells were loaded into the column which was preplaced in the magnetic field of a suitable MACS Separator (Miltenyi Biotec, Germany). The unlabeled cells were collected after flow-through with two times wash processes. The labeled CD26⁺ cells were bound to the column and then flushed out after removing the column from the separator by help of a plunger. (b) Analysis of CD26 expression in the CD26 high-expressing (CD26^high) group and the CD26 low-expressing (CD26^low) group by flow cytometry. Data represented from a minimum of five independent experiments with at least five healthy donor HPBL samples. (c) The dot plots show one typical experiment for analysis of CD26 expression.

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