Colloidal suspensions of copper nanoparticles were synthesized using a developed micro-electrical discharge machining (μ-EDM) method. Average size of the particles dispersed in de-ionized (DI) water is found to be in sub-micron size and that in DI water with stabilizers PolyVinyl Alcohol (PVA) and PolyEthylene Glycol (PEG) is less than 10. nm. The present paper is focused on an exceptional technique using ultrasound which is used for concentration characterization of copper nanofluids in-line. The mass concentration of copper nanoparticles dispersed in suspension media is determined by Urick and Ament's relationship, and the results are found to be 7. wt%, 7. wt% and 6. wt% in Cu-DI water, Cu-DI water. +. PVA and Cu-DI water. +. PEG nanofluids respectively. This study shows that the ultrasonic velocity in Cu-nanofluids increases as compared to the suspension media. However, it was observed that the nanoparticles in suspension media of determined concentrations alter the ultrasonic velocity by affecting the acoustic time of flight. The results demonstrate that due to strong particle-particle interaction conferred by the aggregation of particles, the ultrasonic velocity in the Cu-DI water nanofluid is found to be decreased as compared to the PVA and PEG nanofluid samples. Further, due to the reduction in size of the particles generated and less concentration in DI water. +. PEG mixture, the ultrasonic velocity in the Cu-DI water. +. PEG nanofluid is found to be decreased as compared to that in the PVA nanofluid sample. © 2015 Elsevier B.V.