An automatic microfluidic system for antibiotic susceptibility testing of the present disclosure at least includes a microfluidic chip. The microfluidic chip includes a fluid storage unit, a reaction unit, a pneumatic micro-pumping unit and a plurality of valve units. The fluid storage unit is provided for storing a bacterial suspension, a broth and an antibiotic solution. The reaction unit includes a first reaction chamber and at least two second reaction chambers. The pneumatic micro-pumping unit is adjacently disposed to the fluid storage unit and the reaction unit for selectively, repeatedly and quantitatively transporting the broth, the bacterial suspension and the antibiotic solution to the reaction unit to form a first mixing solution and at least two second mixing solutions. The valve units include a plurality of pneumatic micro-valves and a plurality of valve control air holes for controlling the opening and closing of the pneumatic micro-valves |