Dynamic analysis of blood components is of great importance in understanding cardiovascular diseases and their related diseases, such as myocardial infarction, arrhythmia, atherosclerosis, cardiogenic pulmonary edema, pulmonary embolism, and cerebral embolism. At the same time, it is urgent to break through the continuous heart blood sampling technique in live rats to evaluate the effectiveness of distinctive ethnic medicine therapy. In this study, a blood microdialysis probe was implanted in the right jugular vein of rats in a precise and noninvasive surgical procedure. Cardiac blood samples were then collected at a rate of 2.87 nL/min to 2.98 mL/min by connecting to an online microdialysis sample collection system. Even more momentously, the acquired blood samples can temporarily be stored in microdialysis containers at 4 °C. The microdialysis-based online continuous blood collection program from rat heart has greatly guaranteed the quality of blood samples, advancing and invigorating the scientific rationality of the research on systemic cardiovascular diseases and evaluating ethnomedicine therapy from the perspective of hematology.