Typical LAMP Protocol (M0374)

Incubate the following reaction at 65°C for 30–60 minutes.
Component Final Concentration
10X Isothermal Amplification Buffer II 1X (contains 2 mM MgSO4)
MgSO4 (100 mM) 6 mM (8 mM total)
dNTP Mix (10 mM) 1.4 mM each
FIP/BIP Primers (25X) 1.6 µM
F3/B3 Primers (25X) 0.2 µM
LoopF/B Primers (25X) 0.4 µM
Bst 3.0 DNA Polymerase (8,000 U/ml) 320 U/ml
DNA or RNA Sample > 10 copies or more
Nuclease-free Water to 25 µl
Total Reaction Volume 25 µl

 General Guidelines:
  1. A LAMP Primer Mix can be prepared with all 4 or 6 (with Loop) primers. A 25X Primer Mix should contain: 40 µM FIP, 40 µM BIP, 5 µM F3, 5 µM B3, 10 µM LoopF, 10 µM LoopB in TE or water.
  2. Reactions should be setup on ice. If room temperature setup is desired, use Bst 2.0 WarmStart® DNA Polymerase (NEB #M0538).
  3. If analyzing via agarose gel electrophoresis or other method requiring opening LAMP reaction vessels, setup secondary analysis area and equipment to avoid contamination. 
  4. Running a no-template control is strongly recommended to ensure amplification specificity.
  5. If optimization is desired, try titrating Mg2+ (4–10 mM final) or Bst 3.0 DNA Polymerase (0.04–0.32 U/µl), or changing reaction temperature (50–72°C).