The effect of overnight shipment on signal transduction machinery.
PBMC were stimulated with anti-CD3 (OKT3) + anti-CD28 antibodies for 5 min (A) or with rhIL-6 for 30 min (B) at 37°C in RPMI medium and lysed immediately after stimulation. Post-nuclear lysates were subjected to SDS-PAGE and proteins were transferred onto nitrocellulose membrane. Subsequently, membranes were probed with anti-pERK (A) or with anti-p-STAT3 (Y705) (B) antibody. Equal protein loading was controlled by probing the membranes against β-actin. Results are shown in a form of scanned blots for 2 representative donors out of 4 tested individuals (for both signaling pathways).