In this study, qPCR was also applied to detecting influenza A H3N2 viruses in EBC samples collected by the device. As observed in Figure S3 , H3N2 viruses were detected in the EBC sample collected from subject #3 with a Ct value of 28, while those for subject #1, #2 were shown below the detection limits. In addition, spiking viruses into the samples in general enhanced the overall qPCR signal as observed in Figure S3 . This on the other hand suggests no inhibition or amplification occurred when amplifying H3N2 viruses in EBC samples using qPCR. According to information shown in Table S2 , subject #3 had a fever, but no other information was available at the time of the experiment. In a previous study, it was indicated that use of the RTube for EBC collection had a very low viral detection rate (7%) compared to nasal swabs (46.8%) [20] . Recently, a mask-like sampler was also tested and proved to be useful in detecting viruses using PCR in exhaled breath [23] . It was indicated that airborne virus detection is difficult due to their low concentration and the presence of a wide range of inhibitors, thus optimized molecular biology should be performed to enhance their detection [45] . Although the number of the subjects tested is limited here, the developed method, i.e., EBC collection and qPCR application, was demonstrated successful in detecting viruses from human exhaled breath. This would offer a non-invasive method for diagnosis of respiratory infections by using EBC. In the future, more patients should be tested with the EBC collection device developed here for viral detections.