There is a lack of data on instrument dexterity and interface resistance with respect to the emerging surgical technology of LESS ports. A comparative analysis was conducted to characterize the force to maneuver laparoscopic instruments at various working angles within three commercially available LESS ports. A novel test fixture was created where working angles of the instruments were systematically varied in both the horizontal and sagittal plane within synthetic skin and rigid inserts. Two standard 5-mm laparoscopic graspers and a 10-mm simulated laparoscope were inserted into the trocars of the SILS™, TriPort™ and GelPOINT™ LESS ports. The positions of the laparoscope and grasper (G1) were fixed, while the working instrument’s position (G2) was systematically varied to create a range-of-motion. The static force required to maintain a specific position for G2 was measured using a digital force gauge for that range-of-motion. The resistance created by each LESS port was most noticeable at greater separation angles. The GelPOINT™ provided the least resistance to instrument movement; while the TriPort™ required the greatest amount of force at all angular positions. The 15-mm skin interface yielded lower overall resistance for all ports compared to the 30-mm skin interface. Resistance created by each LESS port increased with greater angular separation. Increased thickness and rigidity of the abdominal wall resulted in greater static forces and reduced instrument range-of-motion for all surgical ports. LESS port design and geometry heavily influenced overall instrument range-of-motion, as well as the resistance found at extreme separation angles. Surgeons should consider the degree of instrument motion required specific to the procedure being performed when selecting a LESS port.