The ranges of the RQ values were calculated by use of the equation: RQ ϭ 2 Ϫ⌬⌬Ct , with ⌬⌬Ct ϩ SD and ⌬⌬Ct Ϫ SDs. The Micro Fluidic Cards detect a 2-fold difference in gene expression at the 99.7% confidence level. The efficiencies of all target and reference amplifications were nearly identical as analyzed by serial dilutions using 2, 1, 0.5, 0.25, and 0.125 ng of calibrator cDNA. When we plotted the log input amount of cDNA vs the ⌬Ct values, the slope was Ͻ0.1. The correlation coefficients were always Ͼ0.95, which is comparable to our previously established method (5 ). We assessed the within-run and day-to-day imprecision of our assay by measuring 4 identical samples in duplicate; the CVs were Ͻ1.6% and Ͻ2.9%, respectively.As can be seen in Table 1, 30 ABC transporters displayed detectable mRNA expression in human primary monocytes, based on ⌬Ct values Ͼ25. Twenty-six genes were induced more than 2-fold during macrophage differentiation, and 12 genes were sensitive to LXR/RXR agonist incubation, with 10 up-regulated ABC transporters and only 2 down-regulated genes. Although the differentiation-dependent and sterol-regulated induction of ABCA1 and ABCG1 is well established (7 ), parallel transcript profiling, using our Human ABC Transporter TaqMan Low-Density Array, revealed several additional differentiation-dependent ABC transporters and novel LXR/RXR-regulated ABC transporters, including ABCB1 (MDR1), ABCB9, ABCB11 (BSEP), ABCC2 (MRP2), ABCC5 (MRP5), ABCD1 (ALD), ABCD4, and ABCG2. Despite the large dynamic range of gene regulation (e.g., a 24.5-fold induction of ABCA1 by 9-cis RA/T0901317), the range of differential transcript expression between 3 different donors was quite narrow (21.9 -27.4). This implies that the interindividual differences in response are much smaller than the differentiation-dependent and lipid-regulated effects, allowing screening processes with a limited number of probands.These findings are particularly relevant for lipidomics and cardiovascular research, as these ABC transporters are novel candidates for lipid disorders and pharmacologic targets for lipid-modulating drugs.In summary, we have developed a Human ABC Transporter TaqMan Low-Density Array based on the TaqMan chemistry, the Micro Fluidic Card, and the 7900HT Sequence Detection System. Compared with standard TaqMan reverse transcription-PCR methods, the Micro Fluidic Card requires less sample material (2 ng instead of 50 ng of total RNA-equivalents per gene), only one-tenth the volume of TaqMan Universal Master Mix, and much less hands-on time. This assay could be a useful tool for monitoring dysregulated ABC transporter mRNA profiles in human lipid disorders and cancer-related multidrug resistance. Furthermore, the pharmacologic and metabolic regulation of ABC transporter gene expression important for drug development could be analyzed in large screening approaches using this Human ABC Transporter TaqMan Low-Density Array.We thank Ernst Arnoldi, Astrid Potratz, and Andrea Geiger for support and discussions, and Manfred H...

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Background: We have previously developed a test for the diagnosis and prognostic assessment of the severe acute respiratory syndrome (SARS) based on the detection of the SARS-coronavirus RNA in serum by real-time quantitative reverse transcriptase polymerase chain reaction (RT-PCR). In this study, we evaluated the feasibility of automating the serum RNA extraction procedure in order to increase the throughput of the assay.