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Pressure pinched injection can introduce well defined shape of sample plugs into the separation channel1.Proper difference in liquid levels among the reservoirs of a microchip is critical for the successful sample injection,but pressurized flow inside the separation channel should be eliminated because it may lead to significant dispersion of the sample plug(Fig.1A).In this work,a simple mathematical model was proposed to calculate the amounts of buffer and sample solutions that should be added to each reservoir to avoid pressurized flow in the separation channel.To verify the model,15 cyclic olefin copolymer microchips were used and the theoretical plate numbers of fluorescein were obtained with different buffer amounts added to the Buffer Waste(BW)reservoir.The maximum theoretical plate number obtained with varying solution volumes in BW(nmax)was compared with the plate number obtained with the predicted volume from the model(npred)and plotted as the relative error in Fig.1B.The results indicate that the optimum volume of solution can be predicted through simple calculation but the solution volume is not the only factor that determines the separation efficiency.Influence of Laplace pressure from meniscus and other factors on the separation performance were also investigated to ensure the efficient separation.