Determining LOD and LOQ for B. cenocepacia

The LOQ is the lowest analyte DNA concentration that provides an acceptable level of precision (i.e., 3/3 replicates amplified), whereas LOD is defined as the concentration of a measurand that is significantly different from a negative control (i.e., at least 2/3 replicates amplified) [28 (link)]. LOD and LOQ were measured through the qPCR and LAMP assays using tenfold serial dilutions of B. cenocepacia AU1054 and B. cenocepacia J2315 genomic DNA. LAMP products were directly analyzed with both the naked eye, and 2% agarose gel electrophoresis. Furthermore, the absorbance in LAMP products was measured using a Synergy MX spectrophotometer from BioTek Instruments, Inc. (Winooski, VT, USA) at 434 and 560 nm [19 (link)]. A reaction was considered positive when the color value was above 0.05 (ΔOD = OD_434nm − OD_560nm; difference in absorbance of samples at 434 (increased absorbance) and 560 nm (decreased absorbance)). Negative reactions (i.e., wells with ΔOD values below 0.05) were analyzed by 2% agarose gel electrophoresis to confirm if a result is a true negative.