Supplemental Data

Full Text
Physiological Regulation of the Immunological Synapse by Agrin
Adil A. Khan, Christian Bose, Lung S. Yam, Mark J. Soloski, Fabio Rupp

Supplementary Material

Supplemental Figure 1. Proliferative responses of resting splenocytes to immobilized CD3 monoclonal antibody (mAb) as a function of CD3 mAb concentration in the presence of saturating amounts of mAbs against agrin m33 and m247 (sqfil), mAb against CD28 (cirfil), mAbs against agrin and CD28 (sq), or nonspecific mAb against HLA (cir). As a measure of lymphocyte proliferation, [3H]thymidine uptake during the last 12 hours of a 3-day incubation was determined. Each point is the average of triplicate cultures, mean ± SEM. Round bottom 96-well plates were coated with 4-fold serial dilutions of mAb against rat CD3. Rat splenocytes (105) in RPMI medium supplemented with 1 mM 2-mercaptoethanol, 10 mM HEPES, 2 mM glutamine, and antibiotics were cultured for 3 days in the presence of a saturating concentration (5 μg/well) of either mAb against agrin m33/m247, mAb against CD28 (50 ng/ml), mAb against agrin m33/m247, and mAb against CD28 or nonspecific mAb against HLA. Cells were pulsed 12 hours with [3H]thymidine (1 μCi/well) and harvested over glass fiber filters.

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