Journal: Clinical and Experimental Immunology

Article Title: CD6 synergistic co-stimulation promoting proinflammatory response is modulated without interfering with the activated leucocyte cell adhesion molecule interaction

doi: 10.1111/j.1365-2249.2010.04235.x

Figure Lengend Snippet: CD6 co-stimulation with activated leucocyte-cell adhesion molecule (ALCAM) induces a distinct gene transcription profile which is amplified further by interleukin (IL)-2. Difference in gene expression profile between 3 × 104 freshly isolated human peripheral blood mononuclear cells (PBMC) stimulated for 72 h with the immobilized anti-CD3 monoclonal antibody (mAb) (0·25 µg/ml) alone (lane 1), with ALCAM-Fc (1 µg/ml) (lane 2), interleukin (IL)-2 (2·5 ng/ml) (lane 4) or both (lane 3) or compared to unstimulated cells (lane 5). (a) Clustering of 15 000 genes or (b) 60 selected genes regulated differentially relative to the unstimulated cells are depicted. (c) Selected genes expression in CD3-activated cells incubated with a soluble CD6 membrane-distal domain specific humanized immunoglobulin (Ig)G1 non-depleting mAb (lane 7) or an irrelevant isotype control mAb (lane 6) at 10 µg/ml, compared to unstimulated cells (lane 8). (d) Colour bar ranging from 0 to 15-fold change relative to control. RNA was extracted by using Trizol reagent protocol. The labelled RNA was hybridized onto the human gene expression array 8 × 15 K. Hybridization was carried out in Agilent's Surehyb Chambers at 65°C for 16 h. The hybridized slides were washed and scanned using Agilent microarray scanner G Model G2565BA at 5 micron resolution.

Article Snippet: We thank Drs H. Iyer and A. Anand for insightful discussions and Drs S. Rao and M. Vasudevan from Genotypic Technology Pvt Limited, Bangalore, India for their support on the gene microarray data analysis.

Techniques: Amplification, Gene Expression, Isolation, Expressing, Incubation, Membrane, Control, Hybridization, Microarray