Nucleic acid amplification test (NAAT) methods are now numerous and varied. In general, they can be categorized into two broad categories:
- target amplification
- signal amplification
The chief difference between these two categories is:
- In target amplification methods, the nucleic acid sequence of interest is geometrically replicated into potentially millions of copies.
- Signal amplification methods do not increase the number of copies of the target sequence in the sample. Large quantities of signal bind to the target sequence to make the signal measurable.
The following table summarizes some of the available techniques. Note: It is intended to provide an overview, not an exhaustive list of all possibilities.
| Target Amplification Methods | Signal Amplification Methods |
| Polymerase chain reaction (PCR) | Probe hybridization assays (eg., Digene; BD GeneOhm™ for C. difficile ) |
| Ligase chain reaction (LCR) | Branched DNA technologies (bDNA); (eg., HIV-1, HCV viral quantification) |
| Transcription mediated amplification (TMA) | Cleavage based signal amplification (eg., Invader®, TB resistance to rifampin) |
| Strand displacement amplification (SDA) | |
| Reverse transcriptase-polymerase chain reaction (RT-PCR): Amplifies RNA targets | |
| Loop-mediated isothermal amplification (LAMP): Targets viral RNA | |
In addition to NAAT, fluorescence in situ hybridization (FISH) techniques employing nucleic acid probes represent another helpful strategy. FISH assays do not involve amplification and are described in detail in a later section.
Although many alternatives have been developed, PCR and PCR-derived techniques remain the most widely used, primarily because of their simplicity and flexibility. Of the target amplification methods, PCR is the principle for many assays. In some ways, the other methods could be considered "variations on the theme." Although it is beyond the scope of this course to describe each technique in detail, the essentials of PCR and RT-PCR will be reviewed.
