Validation of microchip electrophoresis in DNA analysis has been carried out using an Agilent 2100 Bioanalyzer. With a DNA 500 Assay Kit, the reproducibility and accuracy of fragment sizing of a 10 bp DNA ladder have been shown to be satisfactory with the relative standard deviation and the relative error mostly below 1.0 and 5.0% (n = 12), respectively. Both intraday and interday validations of fragment sizing and quantitation have also been performed with a 7500 Assay Kit (n = 48). Although the results of quantitation are not as good as that of sizing, due to the manual introduction of samples and markers into the chip wells, they are still sufficient to carry out further analyses of practical samples. Based on such reliable results, fast analysis of DNA polymorphisms on the human Y-chromosome has been realized with microchip electrophoresis. The total analysis times of three genomic polymorphisms on the Y-chromosome, Y Alu polymorphism, 47z/StuI, and 12f2, are all within 100 s, and the relative standard deviation and relative error of fragment sizes are below 3.5 and 3.7%, respectively. In addition, a mixture of nine DNA markers on the human Y-chromosome related to examine the cause of spermatogenic failure have been separated successfully with the smallest fragment size difference of 7 bp. Our results demonstrate the potential of microchip electrophoresis in polymorphism analysis with the advantages of high speed, good reproducibility, high precision, and high resolution.