Isothermal NAAT
Includes: LAMP, RPA, HDA, NEAR, and related chemistries · Related: PCR · qPCR · CRISPR-based Diagnostics · Syndromic Molecular Panels
1. Principle
Isothermal nucleic acid amplification tests amplify DNA/RNA at a constant temperature using strand-displacement polymerases (± recombinases/helicases), enabling rapid point-of-care assays without full thermocycling.
2. Step-by-Step Procedure (conceptual)
- Extract or use crude lysis compatible with the platform.
- Incubate at fixed temperature (e.g., ~39–65 °C depending on chemistry).
- Detect via turbidity, fluorescence, lateral flow, or CRISPR collateral cleavage readout.
- Interpret with device software / visual cartridge result.
3. Interpretation
- Often qualitative rapid results (15–60 min).
- Pitfalls: Primer design complexity (LAMP uses many primers); multiplexing harder than qPCR; specificity must be tightly validated; still subject to colonization issues.
4. Clinical Use Cases
- Point-of-care strep, influenza, SARS-CoV-2, malaria, TB (platform-dependent).
- Low-infrastructure settings.
- Some hospital rapid instruments use isothermal or near-isothermal engines.
5. Comparison with Other Methods
| Method | Hardware | Multiplex | Quant |
|---|---|---|---|
| Isothermal NAAT | Simple heater / cartridge | Limited–moderate | Usually qualitative |
| qPCR | Thermocycler | Good | Yes |
| CRISPR-based Diagnostics | Often paired with isothermal pre-amp | Emerging | Emerging |
6. Mnemonic / Visual Aid
No cycling, still copying — temperature stays put, enzymes do the unzipping.
Active Recall
- Name two isothermal chemistries.
- Why is LAMP primer design harder than PCR?
- When prefer isothermal over central-lab qPCR?