Sepsis is a medical emergency and it arises from extreme response of the host to an infection. Current diagnosis in sepsis relies on nonspecific clinical signs and culture-based analysis, which is time-consuming. It is critically important for clinicians to follow a protocol to identify sepsis and administer antibiotic therapy without any delay. Sepsis-specific biomarkers are being assessed for early diagnosis and thus improving the outcome of the sepsis patient. Many cellular molecules have been proposed to be sepsis-specific biomarkers. However, these molecules lack specificity and sensitivity. MicroRNA expression in biological fluids, particularly plasma and other tissues is very specific to disease state and are found to be promising diagnostic biomarkers in sepsis. Therefore, it is essential to extract qualitative and sufficient amount of microRNAs from human plasma for the downstream application of two-tailed RT-qPCR method microRNA needed for detection of sepsis patients. The aim of this study was to find optimal volume of plasma required to measure microRNA as sepsis biomarker. The study also included isolating exosomal microRNA from blood samples to check whether blood can be used for extraction. The study was conducted with healthy donor samples and the extraction is performed using Plasma/Serum Exosome Purification (product 58300, Norgen Biotek Corporation, Canada) and RNA Isolation Mini Kit and Total RNA Purification Kit (product 37500, Norgen Biotek Corporation, Canada). The samples were assessed for its quantity and quality by Qubit® and Nanodrop™ technology. Based on the comparison of amount of exosomal microRNA extracted from different plasma volumes, it can be concluded, that increasing volume of plasma may not give higher quantity of microRNA.