Purpose: In this study, through the comparison of the pressure fluctuation and air exchange volume in negative isolation room according to the type of the door and door opening/closing speeds, which is one of the main factors causing the cross contamination of the negative pressure isolation room, establishes standard operating procedures to prevent cross contamination in high risk infectious diseases and isolation room design. Methods: In this study, the air flow each of the room is analyzed using ANASYS CFX CODE for flow analysis. In addition, the grid configuration of the door is constructed by applying Immersed Solid Methods. Results: The pressure fluctuation due to the opening and closing of the hinged door was very large when the moment of the hinged door opened and closed. Especially, at the moment when the door is closed, a pressure reversal phenomenon occurs in which the pressure in the isolation room is larger than the pressure in the anteroom. On the other hand, the pressure fluctuation due to the opening and closing of the sliding door appeared only when the door was closed, but the pressure reversal phenomenon not occurred at the moment when the sliding door was closed, unlike the hinged door. As the opening and closing speed of the hinged door increases, the air exchange volume is increased. However, as the opening and closing speed of the sliding door is decreased, the air exchange volume is increased. Implications: According to the results of this study, it can be concluded that the pressure fluctuation due to the opening and closing of the hinged door is greater than the pressure fluctuation due to the opening and closing of the sliding door. In addition, it can be confirmed that the pressure reversal phenomenon, which may cause to reduce the containment effect in negative pressure isolation room, is caused by the closing of the hinged door. Therefore, it is recommended to install a sliding door to maintain a stable differential pressure in the negative isolation room. Also, as the opening and closing speed of the hinged door is slower and the opening and closing speed of the sliding door is faster, the possibility of cross contamination of the room can be reduced. It is therefore necessary to establish standard operating procedures for negative isolation room for door opening and closing speeds.