Lymph nodes, blood, urine, pelvic abscess, skinĬlearance of bacteremia resolution and closure of draining lymph nodes and pelvic abscessĬlearance of bacteremia resolution of vertebral osteomyelitis and cord compression normalization of liver enzymes To avoid this confounder, the IFN-γ–induced RNA fold-induction for normal PBMCs with normal plasma was set at 100%, and the IFN-γ–induced RNA fold-induction for the same PBMCs with patient plasma was calculated as a percentage of normal. ![]() Because normal PBMCs across individual donors can show large differences in IFN-γ responsiveness, a difference in fold-induction of IFN-γ–induced RNA might reflect differences between normal values and not the actual inhibitory capacity of the plasma. GAPDH was used as a normalization control, and results are expressed as fold-induction over unstimulated. Amplification was performed with Taqman expression assays (Applied Biosystems). RNA (1 μg) was used for reverse transcription by oligo-dT primer (Invitrogen), and the resulting cDNA was amplified by PCR with the use of the ABI 7500 Sequence detector (Applied Biosystems). ![]() ![]() Total RNA was extracted with the RNeasy kit according to the manufacturer's protocols (QIAGEN). For evaluation of gene expression, PBMCs isolated from healthy donors were cultured in complete RPMI 1640 medium containing 10% normal or patient plasma at 37☌ and were unstimulated or treated with IFN-γ at specified concentrations or IFN-α (1000 U/mL) for 3 hours, at which time cells were harvested for RNA isolation.
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