Objective To establish a sensitive droplet digital PCR ( ddPCR) assay for measuring the proviral DNA load in PBMCs (peripheral blood mononuclear cell) and tissues from SIV-infected animals. Methods According to the standard qPCR method , we first optimized the annealing temperature of the droplet dPCR, and then determined the detection range of the ddPCR. This was followed by correlation analysis of ddPCR and qPCR via detection of the 10-fold serially diluted pGEM-SIVgag477. We repeatedly tested 7 DNA samples extracted from PBMCs of SIV-infected monkeys to study the repeatability of the method . Results The annealing temperature of the ddPCR was determined to be 60°C and the detection range was 0. 3~ 3. 96 × 10⁴ copies/ μL. The correlation coefficient of ddPCR and qPCR was in the range of 0. 9981. Conclusions We established an absolute quantitative method for SIV DNA analysis based on droplet dPCR. This method can be used to accurately quantify the proviral DNA load and represents an effective technique for the measurement of viral reservoirs in SIV research.