Circulating tumor DNA (ctDNA) has been demonstrated as the most promising biomarker for non-invasive assessment of cancer. However, since circulating DNA is a challenging analyte due to little amount in blood. Furthermore, less than 1% is tumor-derived DNA as well as many variations can be found in several hundreds of tumor DNAs even for one cancer. Therefore, integrated system for ctDNA analysis from stable nucleic acids extraction to accurate multiple detection is highly required in cancer management. Here, we present a circulating nucleic acids extraction system under vacuum with immiscible solvent, closed-loop amplification in microfluidic PCR for amplification of target DNA and lab-made surface plasmon resonance (SPR) assay for detecting epidermal growth factor receptor (EGFR) mutations. Due to characteristics of SPRi (SPR image), multiple mutations were monitored in real-time within an hour. This study as a proof of concept can be further expanded into automated sample to answer system for high degree of multiplexing detection of major and known ctDNAs, which could provide a solution for clinical unmet needs in cancer treatment and early detection.