Self-adhesive resin cement (G- CEM, USA) was used for fixing of metal tube to the abutment. A customized rigid metal mounting jig was designed, as a holding device, to ensure solid fixation of the sample while recording the measures.

Abutment screws were tightened in all the groups to 30 Ncm using a digital torque gauge (HTG2 - 200Nc, IMADA, Toyohashi, Japan) according to the manufacturer’s recommendations to ensure an accurate application of reproducible force to each abutment screw every time for standardization. Ten minutes after first torque application, all screws were retightened to the same tightening torque (30 Ncm). Ten minutes later, RTV before loading was measured and recorded.

At each group, after measuring the initial removal torque value, the abutment screw was tightened again to the recommended torque value (30 Ncm).

The acrylic resin block was firmly mounted in a holder of the lower fixed compartment of a computer-controlled universal testing machine (Model 3345; Instron Industrial Products, Norwood, MA, USA) for 100,000 cycles of eccentric dynamic cyclic loading.

Dynamic cyclic loading was performed with a metallic rod with a round tip which was attached to the upper movable compartment of the machine, under a load of 130 N with contact time between the rod and the metal tube 0.2 s at a rate of 1 Hz which simulates the tooth contact duration of each masticatory cycle. The load was perpendicular to the metal tube and 5 mm away from the center axis of the implant (Fig. 4). After that, the acrylic resin block with fixture and abutment was transferred again to the metal jig to measure RTV after loading.

Screw loosening of each assembly (implant–abutment) was analyzed by measuring RTV before and after dynamic cyclic load by using the digital torque gauge. RTL can be an indicator of how much loosening takes place.

Each RTL ratio was calculated using the following formula: