40 w led light

HuNoV detection in oysters using PMA pretreatment was performed using the method described in Jeong et al. [27 (link)]. For dye treatment, treated virus samples (each 200 µL) were immediately mixed with 200 µM PMA (Biotium, Hayward, CA, USA). These mixtures were left in the dark at room temperature for 10 min to allow dye penetration. After that, the samples were exposed to 40 W LED light (Dynebio, Seongnam, Korea) at a 460 nm wavelength at room temperature for 20 min to photoactivate both dyes. HuNoV with damaged capsids and HuNoV with intact capsids were distinguished for HuNoV infectivity through PMA pretreatment. HuNoV infection samples without PMA pretreatment were set as control samples.

To allow for dye penetration, the virus-treated samples were immediately combined with 200 µM PMA (Biotium, Hayward, CA, USA) and incubated in the dark for 5 min at room temperature. Then, samples were photoactivated for 10 min on each side with 40 W LED light (Dynebio, Seongnam, Republic of Korea) at 460 nm wavelength at room temperature and distance was 3 mm from the samples [1 (link)]. A control group was included in the study that was not treated with PMA or exposed to halogen light to examine if the dye treatment interfered with viral identification. Sodium lauroyl sarcosinate (Sigma-Aldrich, St. Louis, MO, USA) was used at 1.0% (w/v) to optimize the sarkosyl concentration and to examine its effect on HuNoV [18 (link)]. Sarkosyl solution was added to the PMA mixture at the same time as detergent treatment. PMA + sarkosyl-treated samples were incubated for 10 min at room temperature in the dark. An untreated control was included and unexposed to LED light to examine the ability of PMA + sarkosyl treatment to interfere with HuNoV detection [9 (link),17 (link),18 (link)]. Finally, before RT-qPCR experiments for discriminative detection of possibly infectious and noninfectious HuNoV viral particles, viral samples were exposed to the optimized PMA and sarkosyl pretreatments.