A novel strategy integrating methodological calibration and analysis on board on a planar first-generation microfluidics system for the determination of total isoflavones in soy samples is proposed. The analytical strategy is conceptually proposed and successfully demonstrated on the basis of (i) the microchip design (with the possibility to use both reservoirs), (ii) the analytical characteristics of the developed method (statically zero intercept and excellent robustness between calibration slopes, RSDs < 5%), (iii) the irreversible electrochemical behaviour of isoflavone oxidation (no significant electrode fouling effect was observed between calibration and analysis runs) and (iv) the inherent versatility of the electrochemical end-channel configurations (possibility of use different pumping and detection media). Repeatability obtained in both standard (calibration) and real soy samples (analysis) with values of RSD less than 1% for the migration times indicated the stability of electroosmotic flow (EOF) during both integrated operations. The accuracy (an error of less than 6%) is demonstrated for the first time in these microsystems using a documented secondary standard from the Drug Master File (SW/1211/03) as reference material. Ultra fast calibration and analysis of total isoflavones in soy samples was integrated successfully employing 60 s each; enhancing notably the analytical performance of these microdevices with an important decrease in overall analysis times (less than 120 s) and with an increase in accuracy by a factor of 3.