The Maxwell® CSC Instrument (Figure 1) and Maxwell® CSC Blood DNA Kit are the first products from Promega developed and manufactured under current Good Manufacturing Practices (cGMP). The products are intended for use as clinical sample concentrators for in vitro diagnostic use in the US market. The Maxwell® CSC Instrument offers a variety of features specifically tailored for the clinical laboratory, including availability of Installation Qualification (IQ)/Operational Qualification (OQ) services, multiple tiers of service and support packages, 21 CFR Part 11-compliant software, an integrated touch-screen tablet providing an easy-to-use graphical user interface, bar code reading and sample tracking capabilities, administrator-level instrument-specific and workflow-specific options, and reporting functionality. The new software platform for operating the Maxwell® CSC Instrument has a tailored user interface that suits the needs of the clinical laboratory. Selecting a protocol and keeping track of kit lot number and expiration date are as simple as scanning a bar code on the Maxwell® CSC reagent kit. Purification setup instructions within the software require users to confirm that they have performed all of the steps necessary for a sample processing run.
The first reagent kit available for the Maxwell® CSC Instrument is the Maxwell® CSC Blood DNA Kit. This new kit has undergone extensive testing to ensure reliable and reproducible DNA purification from whole blood samples when it is used with the Maxwell® CSC Instrument. We conducted cross-contamination studies to demonstrate that no detectable sample contamination occurs during processing, and inhibitor studies to show that amplification inhibitors do not copurify with the eluted DNA. This article describes some of the design verification and validation experiments performed during product development.
We designed the Maxwell® CSC Blood DNA Kit to purify DNA from human whole blood with a normal range of white blood cells (WBC; 4 × 106–10 × 106WBC/ml of blood) in the anticoagulant blood collection tubes most commonly used in clinics: EDTA, heparin and sodium citrate. Figure 2 shows the yield and average purity of nucleic acid purified from eight replicate 300μl blood samples from a single donor collected using different blood anticoagulant tubes and purified using the Maxwell® CSC Blood DNA Kit. Together, these data demonstrate that the Maxwell® CSC Blood DNA Kit provides reproducible yield and purity from samples collected in the common anticoagulant blood tubes tested.
We designed the Maxwell® CSC Blood DNA Kit to work with blood sample volumes of 50–300µl. Figure 3 shows the linearity of DNA yield across this range. In addition, we tested the ability of the Maxwell® CSC Blood DNA Kit to purify DNA from blood samples stored up to 72 hours at ambient temperature, up to 7 days at 4°C, or frozen at –80°C and thawed at room temperature with mixing before extraction. We found that blood stored under these three conditions produced comparable DNA yields and purities when processed with the Maxwell® CSC Blood DNA Kit (Figure 4).
Multiple analysts are often responsible for processing blood samples within a single laboratory. For this reason, we evaluated the consistency of DNA extraction over multiple days, multiple instruments and by multiple operators using blood from a single donor. Over a five-day period, a single user processed eight replicates per day of the same frozen blood sample to assess the consistency of extraction. The results of this multiple-day testing were comparable for yield and purity (Figure 5). We also tested the consistency of extraction performance when using three different Maxwell® CSC Instruments, and in a separate experiment, we assessed DNA extraction consistency among three different users. Again we found reproducible yield and purity for each of the three instruments tested (Figure 6) as well as each of the three analysts (data not shown). Together these results indicate that the Maxwell® CSC System provides consistent performance over multiple days, multiple instruments and multiple operators.
Whether samples are processed manually or with an automated system, one of the main concerns of any laboratory is ensuring that there is no cross-contamination between samples. We assessed cross-contamination by dispensing water and blood samples in alternating Maxwell® CSC Blood DNA Kit cartridges that were processed in the Maxwell® CSC Instrument. The eluted DNA was analyzed using real-time PCR to quantitate picogram amounts of human genomic DNA. Triplicate amplifications of each eluate from the water sample cartridges were tested and showed no detectable human genomic DNA contamination. This indicated that there was no cross-contamination of samples during processing with the Maxwell® CSC Instrument (Table 1).
Each lot of Maxwell® CSC Blood DNA Kits is functionally tested during quality control and must meet a stringent set of performance requirements. During testing, 15 replicate 300µl samples of whole blood and one water sample are processed in 16 Maxwell® CSC Blood DNA Kit cartridges with the kit reagents provided for that lot. The DNA is purified on a Maxwell® CSC Instrument with a final elution volume of 50μl. The eluates are assessed using absorbance, and the minimum acceptable concentration is 80ng/µl with purity values required to be A260/A280 > 1.7 and A260/A230 > 1.5 for each replicate.
During design verification and validation testing, we assessed the performance of the Maxwell® CSC Blood DNA Kit and the Maxwell® CSC Instrument. The Maxwell® CSC Blood DNA Kit will typically yield 150–400ng/μl of DNA with absorbance purity ratios of A260/A280 = 1.85–1.95 and A260/A230 = 1.9–2.2 from 300µl of whole blood (4 × 106–10 × 106WBC/ml of blood) and an elution volume of 50μl.
With a small footprint, easy-to-use interface, sample-tracking and reporting features, cGMP manufacture and 21 CFR Part 11-compliant software, the Maxwell® CSC Instrument is an ideal extraction robot for the clinical laboratory. The Maxwell® CSC Blood DNA Kit offers laboratories a reliable chemistry that performs consistently across a range of sample and elution volumes, works with common anticoagulant blood collection tube types and provides DNA samples free from cross-contamination with minimal instrument-to-instrument and day-to-day variability. Together, the Maxwell® CSC Blood DNA Kit and Maxwell® CSC Instrument constitute a reliable and consistent platform for extracting genomic DNA from human whole blood samples for use in amplification-based diagnostic assays. While the Maxwell® CSC Blood DNA Kit is our first cGMP-compliant reagent kit, additional Maxwell® CSC reagent kits specifically designed to address the needs of the clinical laboratory are being developed.