A total of 48 water samples were processed on the BioRobot MDx workstation using the PAXgene Blood RNA MDx Kit. The eluates were pooled and added to quantitative, real-time RT-PCR assays at the percentage indicated. Each assay was performed in quadruplicate using the same amount of RNA from HeLa cells, with primers and probe for β-actin. [A]CT values are given (mean ± standard deviation), with the difference between the mean CT values and the mean CT value for the 0% reactions (ΔCT). None of the reactions were inhibited (ΔCT ≥0.5). [B]Amplification curves for all 20 assays show no detectable inhibition of RT-PCR.
Following collection of blood samples (2.5 ml) in PAXgene Blood RNA Tubes, the simple purification procedure begins with a centrifugation step to pellet the contents of each PAXgene Blood RNA Tube. Proteinase K and an optimized incubation buffer are added to digest proteins. The tubes are then transferred to the BioRobot MDx or BioRobot Universal System for automated RNA purification.
Blood was collected in duplicate from 3 donors into PAXgene Blood RNA Tubes. RNA was purified using the PAXgene Blood RNA MDx Kit on the BioRobot MDx instrument. RNA was analyzed with an Agilent 2100 Bioanalyzer. The RNA integrity number (RIN), indicated on each scan, shows the high quality of the purified RNA. [A] Analysis of 2 samples from donor A. [B] Analysis of 2 samples from donor B. [C] Analysis of 2 samples from donor C.
RNA was purified from 384 blood samples collected in PAXgene Blood RNA Tubes and processed in 4 runs on the BioRobot MDx using the PAXgene Blood RNA MDx Kit. Genomic DNA was quantified using a genomic-DNA-specific real-time TaqMan PCR assay with primers and probe for the β-actin gene. The amount of DNA present is given as a percentage of the total nucleic acid content.
Blood was collected in fivefold replicates from 6 donors into PAXgene Blood RNA Tubes. RNA was purified using the PAXgene Blood RNA MDx Kit on 3 different BioRobot MDx workstations (MDx 1, MDx 2, MDx 3) and manually using the PAXgene 96 Blood RNA Kit (PAX 96). Purified RNA was analyzed using quantitative, real-time RT-PCR, with primers and probe for the c-fos transcript. All values were well within the ±3x total precision of the assay (2.34 CT), indicated by the horizontal lines.