Molecular diagnostics: a revolution in progress.

Molecular diagnostics now provides most laboratory tests in infectious disease and genetics, and an increasing number in oncology. Massively parallel methods allow sequencing of entire genomes, and sequence databases within and between species provide the information necessary to develop sensitive and specific diagnostic assays. Indeed, most microorganisms today are classified on the basis of sequence rather than phenotype. If a microbial sequence is present where it should not be, a presumptive diagnosis is made. Modern genetics, empowered by the 3.08 billion–base book of the human genome, has advanced beyond singlegene disorders to gene families, exomes, transcriptomes, and ultimately, to the complete genome. Severe challenges in data processing and interpretation remain, but the enormity and potential closure of the undertaking is inspiring. New classes of nucleic acid biomarkers, including long noncoding RNAs and microRNAs, provide a deep well for correlation to disease. If a patient’s sequence variation is consistently associated with disease, a genetic diagnosis is made. Similarly, cancer is caused by genome sequence variations. These variations are typically widespread, with myriad copy number and sequence changes throughout the genome. Some of these variants identify molecular pathways that can be blocked or enhanced by drugs that affect tumor growth. Molecular diagnostics is empowered by continual advancements in technology. Applications of massively parallel sequencing have exploded, affecting all areas of medicine, including molecular diagnostics. The polymerase chain reaction has evolved from gel analysis to real-time PCR to, most recently, digital PCR, where individual targets are amplified in picoliter to nanoliter volumes and statistics are used to calculate target concentrations instead of standard curves or internal controls. Genome-wide studies are no longer considered fishing expeditions, and disease association studies that use expression, single nucleotide polymorphism, and copy number microarrays have identified useful markers that have been reduced to practical molecular tests. Big data needs have resulted in new bioinformatics tools that continue to evolve rapidly. But wait, just what is molecular diagnostics? The words, “molecular” and “diagnostics” do not appear together in the scientific literature until the mid1980s (Fig. 1). Since 1985 (around the time that PCR appeared), these words increasingly cohabit scientific abstracts at an exponential rate, doubling every 6–7 years. “Molecular” came to mean DNA or RNA, unfairly excluding proteins, electrolytes, small molecules, and other polymers. Substantial growth of molecular diagnostics in clinical laboratories was deferred over 10 years, taking off around 1997 in parallel with the commercial availability of real-time PCR instruments (Fig. 2). The number of tests offered by clinical laboratories rose linearly until 2006 and then assumed a faster, but still linear, rate of growth through 2014. The volume of testing performed over time was 1 Department of Chemical Pathology, The Chinese University of Hong Kong, Hong Kong SAR, China; 2 Department of Pathology, University of Utah School of Medicine, Salt Lake City, UT; 3 ARUP Laboratories, Salt Lake City, UT. * Address correspondence to this author at: Department of Pathology, University of Utah Medical School, 50 NMedical Dr, Salt Lake City, Utah 84132. Fax 801-581-6001; e-mail [email protected]. Received October 31, 2014; accepted November 3, 2014. DOI: 10.1373/clinchem.2014.233361 © 2014 American Association for Clinical Chemistry Carl T. Wittwer Y.M. Dennis Lo