Stability of Loop-Mediated Isothermal Amplification (LAMP) reagents and its amplification efficiency on crude trypanosome DNA templates.
This study evaluated the stability of LAMP reagents when stored at 25 degrees C and 37 degrees C, and also assessed its detection efficiency on different DNA template preparations. Accordingly, LAMP using reagents stored at 25 degrees C and 37 degrees C amplified DNA of in vitro cultured T. b. brucei (GUTat 3.1) from day 1 to day 15 of reagent storage. There were no significant differences (P>0.05) in detection sensitivity of LAMP among the reagents stored at 25 degrees C, 37 degrees C and -20 degrees C (recommended storage temperature). LAMP using the reagents stored at above-mentioned temperatures amplified serially diluted DNAs (genomic DNA extracted by phenol-chloroform method, FTA card and hemolysed blood) of T. b. gambiense (IL2343) with high sensitivity. Reactions were conducted on the reagents stored from 1 day to 30 days. LAMP detection sensitivity was poor when fresh blood as DNA template was added directly into reactive solution. Results of this study demonstrated that LAMP has the potential to be used in field conditions for diagnosis of trypanosome infections without being affected by ambient temperatures of tropical and sub-tropical countries where trypanosomosis is endemic.
National Research Center for Protozoan Diseases, Obihiro University of Agriculture and Veterinary Medicine, Japan.
Oriel M M
OM
Thekisoe
Raoul S B
RS
Bazie
Andrea M
AM
Coronel-Servian
Chihiro
C
Sugimoto
Shin-Ichiro
S
Kawazu
Noboru
N
Inoue
D000818:Animals; D002417:Cattle; D016054:DNA, Protozoan / Q000737:chemistry / Q000235:genetics; D021141:Nucleic Acid Amplification Techniques / Q000379:methods / Q000662:veterinary*; D012680:Sensitivity and Specificity; D014346:Trypanosoma brucei brucei / Q000235:genetics* / Q000302:isolation & purification*; D014354:Trypanosomiasis, Bovine / Q000821:virology*
D016054:DNA, Protozoan