Over the years, polymerase chain reaction (PCR) has evolved into a readily automated, high throughput quantitative technology. Real-time quantitative PCR (qPCR) has become the industry standard for the detection and quantification of nucleic acids for multiple application, including quantification of RNA levels. But a lack of consensus among researchers on how to best perform and interpret qPCR experiments presents a major hurdle for advancement of the technology. This problem is exacerbated by insufficient experimental detail in published work, which impedes the ability of others to accurately evaluate or replicate reported results.
In extreme instances, incongruous pre-assay conditions, poor assay design and subjective data analysis methods have led to the publication of irrelevant or misleading data. The original 2002 paper on Measles, Mumps and Rubella (MMR) provided evidence that supported a link between the MMR vaccine and autism. Later examination established the original conclusions were based on flawed real-time PCR data. Subsequent publications have convincingly demonstrated there is no plausible link between the vaccine and autism, but the debate and doubt continues outside of the scientific community and MMR vaccination rates have dropped in several countries.
Accurate qPCR testing can help companies avoid pursuing the wrong compound or target. Providing sufficiently detailed reports of experimental design in early stage drug discovery research can potentially save millions of dollars by not wasting man-hours or reagents. The Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) standards can help companies and researchers avoid these drug development pitfalls.
MIQE’s goal is to restructure the current approach to qPCR data reporting into a consistent format that encourages detailed auditing of experimental processes, analysis and results whereby the technical quality of the work and reliability of the conclusions can be evaluated. Implementing these guidelines is imperative in continuing the growth of qPCR into an accurate and reliable nucleic acid quantification technology.
MIQE promotes the careful examination of qPCR results to enhance integrity of submissions to peer-reviewed journals, consistency between laboratories and experimental transparency. Adoption and compliance of these guidelines will help researchers more accurately report data and replicate experiments, ultimately saving time and money.
When it comes to providing scientists with the tools to be MIQE-compliant, Bio-Rad Laboratories, Inc. is paving the way. Recently, Bio-Rad entered into an arrangement with Biogazelle to exclusively distribute the qbasePLUS data analysis software with its CFX96 and CFX384 real-time PCR detection systems, a program that enables users to annotate their experiments with MIQE-compliant experimental details. The qbasePLUS software in combination with these real-time PCR systems provides customers with improved accuracy in their qPCR experiments and speeds up standardized data analysis.
qbasePLUS software allows for the elimination of erroneous data, normalization to remove sample-specific non-biologic variation, and inter-run calibration, which can remove the technical variation between samples analyzed in different runs. By enhancing the reliability of the qPCR data, qbasePLUS conforms to MIQE.
Using the CFX96 and CFX384 real-time detection systems with qbasePLUS software accelerates research through automated, fast calculations and direct import of data from the system to the software. The combined solution provides researchers with the flexibility to handle both small and large experiments as well as combine data from different experiments using qbasePLUS software as the repository database.
As the industry continues to recognize the importance of the MIQE guidelines it will become increasingly necessary to ensure that laboratories and corporations are following them. Simplifying compliance facilitates not only drug discovery and development, but basic biological research. Investing in a qPCR system and failing to follow guidelines that will ensure that quality data is being produced may result in a wasted investment.
It can be anticipated that additional qPCR vendors will soon offer solutions to assist researchers in maintaining MIQE compliance. Top instrument makers and reagent vendors are supporting the thought leaders working to expand MIQE awareness and compliance. Bio-Rad has trained sales and support staff to help researchers comply with the guidelines. They share this training through educational meetings focused on MIQE.
Quality data is paramount for the successful development and potential approval of a drug candidate. Following MIQE will help strengthen the quality qPCR data needed to move a candidate forward. Plus, it may avoid the devastation of finding out the focus was on the wrong target or having a drug candidate fail.
Richard Kurtz, PhD is senior marketing manager at Bio-Rad Laboratories. Prior to joining Bio-Rad Kurtz served as field applications manager for MJ Research Incorporated. Kurtz earned his PhD from Northwestern University and a BA in Molecular Biology from Colgate University.