Summary and Future Direction

In the past decade, probe amplification technologies have advanced significantly, from the initial description of Q-beta replicase amplification in 1986 (Chu et al., 1986) to the most recently introduced RAM (Zhang et al., 1998). It is expected that more probe amplification methods will be invented in the next 10 years, and the applications of the current probe amplification methods will become more diversified. Homogeneous and real-time monitoring of amplification will be devised to probe amplification technologies to reduce detection time and improve quantification capability of the assay. Additional technologies will be developed to be used for the detection of RNA, DNA, and protein (antigen/antibody) on a single platform, which will further enhance the detection sensitivity and specificity. Finally, the applications of these technologies will become broader as the fields of genomics, proteomics, and pharmacogenomics advance. Therefore, a technology that offers in situ detection and amplification, microarray, immunoassay, real-time monitoring, whole-genome amplification, and SNP detection will be more favorable. However, no single technology can meet all of these requirements, and possible combination of these technologies may be the answer. Also, PCR, the dominant amplification technology, cannot fulfill all these needs, and ample room is available for probe amplification technology to grow.

On the other hand, the instrumentation for probe amplification will change significantly in the next 10 years. Fluorescence-based real-time detection instrument will be widely used in the diagnostic laboratory, which will certainly improve throughput. Miniaturized microfluidic assay format will soon be available in the clinical laboratory, which will significantly reduce sample volume. Automation and miniaturization of the instrument will make molecular diagnosis at a doctor's office and at the bedside possible. It is expected that the array-based assay and instrument will be significantly improved, and the cost will be reduced to an affordable level. Given the advantages of probe amplification (isothermal, multiplex, on-chip amplification, etc.), probe-based amplification could be easily adapted in these formats and will become the dominant technologies in clinical diagnostic applications.

However, most described probe amplification technologies are still at the early stage of development. Most publications only demonstrated the feasibility in clinical diagnosis, and their clinical performance has not yet been demonstrated in large clinical trials. It is anticipated that some of these technologies may not meet the clinical diagnostic requirements and will consequently be lost in market competition. For example, Q-beta replicase technology did not reach the clinical laboratory even after an initial favorable clinical trial, and the LCx assay (LCR technology) for Chlamydia was voluntarily withdrawn by Abbott in 2003 due to significant reproducibility problems (Gronowski et al., 2000). Therefore, it is expected that more changes (exciting or disappointing) will happen in the field of probe-based amplification technologies in the next 10 years.

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