FAQ: Amplification occurred in my NTC sample(s) following isothermal incubation. What happened?

Amplification in the non-template control within 30 minutes may indicate cross-contamination during reaction set up or a systemic issue. 

Some primer sets are more susceptible to non-specific amplification than others. We recommend evaluating at least two LAMP primer sets for any given target. In addition, some reaction formats or workflows may be more prone to non-specific amplification with particular primer sets such as: 

  • Large reaction volumes in small vessels (e.g., 20 uL in 384-well plates)
  • Low reaction temperatures (e.g., 60°C instead of 65°C)

If using a real-time thermocycler, a melt or denaturation curve can be included after the LAMP incubation to distinguish between spurious amplification and cross-contamination since each LAMP amplicon will produce a unique melt profile. 

If the NTC reaction is positive upon repeat testing and/or workflow changes, replace all reagent stocks and clean workspace with an acceptable surface decontaminate such as 10% bleach (1:10 dilution of commercial 5.25-6.0% sodium hypochlorite).