LAMP Reaction Orange Before Amplification Starts

Symptom

The colorimetric LAMP reaction appears orange instead of pink/magenta prior to incubation at 65°C, indicating premature pH drop that compromises amplification.

1. 1 Nucleic acid sample contains acidic extraction buffer or contaminants that lower reaction pH below optimal ~8.0
2. 2 Sample input volume is incompatible with buffering capacity of the master mix
3. 3 Repeated exposure of colorimetric LAMP master mix to atmosphere causes CO₂ absorption and gradual pH drop
4. 4 Extended time between master mix thawing and reaction setup allows pH drift

1. 1 Dilute sample in nuclease-free water prior to addition to the reaction to reduce inhibitor concentration
2. 2 Adjust sample pH to ~8.0 using Tris or HEPES buffer before adding to the LAMP master mix
3. 3 Minimize master mix air exposure by immediately closing the cap after each use
4. 4 Store opened colorimetric LAMP master mix at -20°C in the original vial provided
5. 5 Prepare reactions immediately after thawing master mix components