The BioRobot MDx is the latest addition to the range of BioRobot workstations and is designed to meet the needs of scientists requiring standardized protocols in environments such as clinical testing laboratories.



Benefits of the BioRobot MDx:

• Standardized nucleic acid purification and reaction setup - proven QIAamp technology provides high-quality nucleic acids
  
• Purify genomic and viral nucleic acids - genomic DNA from fresh or frozen whole blood, and viral RNA and DNA from serum and plasma
  
• Walkaway automation automated vacuum system eliminates manual centrifugation steps
  
• High process reliability bar- or colorcoded reagent vessels and accessories plus a detailed loading check ensure smooth sample processing
  
• Intuitive software QIAsoft MDx Operating System provides a user-friendly interface

Efficient protocols

Optimized protocols for nucleic acid purification and reaction setup are provided with the BioRobot MDx. The QIAamp Virus BioRobot MDx Kit and the QIAamp DNA Blood BioRobot MDx Kit are used for purification of viral nucleic acids and genomic DNA, respectively. The protocols offer rapid and efficient purification, requiring only 2.5 hours preparation time for 96 samples, including a load check of 15 minutes. Table 1 shows examples of applications using these kits. Use of the QIAamp DNA Blood BioRobot MDx Kit results in highly pure nucleic acids that perform well in sensitive downstream assays. The QIAamp Virus BioRobot MDx Kit provides efficient purification of low amounts of viral nucleic acids. Protocols are also supplied with the instrument for reaction setup using commercially available amplification systems. Additional reaction setups can be programmed by our application specialists.

Table 1. High precision using the QIAamp Virus BioRobot MDx Kit



Plasma was spiked with armored RNA molecules to mimic RNA viruses (25,000 copies per prep) and processed using the QIAamp Virus BioRobot MDx protocol in replicates of 96 on two separate instruments. An internal control RNA was added to the lysis buffer. The eluted RNA was amplified using a TaqMan RT-PCR system. Mean CT value, standard deviation, and %CV were calculated for both the mimic virus and the internal control. The precision of the assays was <1%, providing highly reproducible data. (Data generated by QIAGEN R&D department.)

Effortless Sample Tracking



Figure 1 Sample tracking system.

High reliability load checks, full data tracking, and process documentation

Up to 12 types of primary sample tube can be used in each run. User-friendly colorcoding of tip trays and bar- or color-coded, individually shaped reagent vessels greatly facilitate worktable setup. A detailed load check enhances process reliability by ensuring that all reagents needed are present, and that buffer volumes are sufficient for the run. Sample tubes and tube holders are labeled with bar codes, eliminating the risk of sample mix-up. The data tracking process includes a sample transfer check, which ensures that samples have been transferred correctly, and flags samples in which an error was detected.
All information relevant to the run can be exported to local or external networks.

Enhanced user safety

The system can be locked during the run, preventing access to the worktable, and primary sample tubes are covered during processing. These features reduce the users exposure to potentially infectious material and greatly reduce cross contamination. A conveniently placed tip-tray drawer ensures that users do not need to lean over the worktable to replace tips, further improving safety.

Conclusions

The new BioRobot MDx workstation provides a robust and efficient system for automated purification of nucleic acids. Walkaway automation, process documentation, and intuitive software greatly facilitate a range of assays in clinical laboratories.

Principle of siRNA-mediated RNA interference siRNA-Mediated RNAi
The application of RNAi technology to mammalian cells has revolutionized the field of functional genomics. Use of siRNA allows targeted inhibition of gene expression using double-stranded RNA (dsRNA) that carries the same sequence as mRNA transcribed from the target gene. In mammalian cell systems, these double-stranded siRNA oligos can be used to induce gene silencing.

Targeting a specific gene by RNAi requires a double-stranded siRNA containing the sense and antisense sequence of mRNA transcribed from the target gene. QIAGEN uses patented TOM-amidites to synthesize high-quality, highpurity, 21-nucleotide sense and antisense RNA oligonucleotides. In mammalian cells, siRNA must be delivered to the cytoplasm using an RNA transfection reagent. TransMessenger Transfection Reagent from QIAGEN efficiently delivers siRNA to cells for successful gene knock down (Figures 1 and 2).

Once inside the cell, current models suggest that siRNA interacts with endogenous cellular proteins to form a proteinRNA complex known as the RNA induced silencing complex (RISC). Due to the sequence homology between the siRNA and the target mRNA, the RISC specifically targets the mRNA for enzymatic degradation (see flowchart).
Efficient Gene Silencing Using QIAGEN siRNA and TransMessenger Reagent Efficient, Targeted Gene Silencing Figure 1 Western blot of extracts from transfected HeLa-S3 cells probed with lamin A/C monoclonal primary antibodies. Cells were transfected using the indicated reagents. siRNA: short interfering RNA duplex targeting lamin A/C at position 608630 relative to the start codon; TM: TransMessenger Transfection Reagent; ODN: Scrambled 24mer oligodeoxynucleotide (control). Figure 2 Relative expression levels of lamin A/C after siRNA transfection. The value for the mock transfection was set as 1. Data were normalized to the expression level of GAPDH (internal control), which was quantified by real-time RT-PCR using OmniscriptReverse Transcriptase and the QuantiTect SYBR Green PCR Kit. Expression levels were calculated using a standard curve generated using different amounts of HeLa S3 cDNA. Data obtained under the indicated transfection conditions are shown as relative, normalized expression levels. Total RNA was prepared using the RNeasy Mini Kit and the QIAShredder homogenizer. High-quality siRNA for efficient gene silencing

QIAGEN offers two grades of custom siRNA; ultrapure siRNA purified by ion-exchange HPLC (>97% pure), and crude siRNA (8085% pure). In addition, a number of library siRNA duplexes (>97% pure) are available for silencing common target genes. All siRNA products are delivered annealed,* desalted, deprotected, and ready to use. A detailed certificate of analysis, including HPLC analysis, is included with each order. All QIAGEN siRNA is supplied with a sterile resuspension buffer and is suitable for use in transfection procedures.

* Non-annealed siRNA is available on request.
HPLC trace of the siRNA duplex is provided. If non-annealed siRNA is requested, two individual chromatograms of the single strands are provided.




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