Blood cell manipulation in microdevices is an interesting task for the separation of particles, by their size, density, or to remove them from the buffer, in which they are suspended, for further analysis, and more. This study highlights the cell-free area (CFA) widening based on experimental results of red blood cell (RBC) flow, suspended in a microfluidic device, while temperature and flow rate incrementally modify RBC response within the microflow. Studies of human red blood cell flow, at a concentration of 20%, suspended in its autologous plasma and phosphate-buffered saline (PBS) buffer, were carried out at a wide flow rate, varying between 10 and 230 μL/min and a temperature range of 23 °C to 50 °C. The plotted measures show an increment in a CFA near the channel wall due to cell flow inertia after a constricted channel, which becomes more significant as temperature and flow rate increase. The temperature increment widened the CFA up to three times. In comparison, flow rate increment increased the CFA up to 20 times in PBS and 11 times in plasma.