An analysis of biosignal and performance data collected during driving has increasingly employed in research to explore a human-vehicle interface design for better safety and comfort. The present study developed a protocol and a system to effectively analyze biosignal and driving performance measurements in various driving conditions. Electrocardiogram (ECG), respiration rate (RR), and skin conductance level (SCL) were selected for biosignal analysis in the study. A data processing and analysis protocol was established based on a comprehensive review of related literature. Then, the established analysis protocol was implemented to a computerized system so that immense data of biosignal and driving performance can be analyzed with ease, efficiency, and effectiveness for an individual and/or a group of individuals of interest. The developed analysis system would be of use to examine the effects of driving conditions to cognitive workload and driving performance.