The mechanical properties of gel sheets were measured at a stretching speed of 1 mm/min with a desktop universal testing machine (EZ test; Shimadzu, Kyoto, Japan), of which maximum load cell capacity was 500 N under standard ambient conditions at 25 ± 3 °C and 50 ± 25% RH. The samples were gripped by clamps at each end (using slip-proof rubber sheets to prevent slippage) such that the initial apparent gauge length (the distance between clamp faces) was set to 10 mm for all the samples tested.

Young’s modulus, maximum tensile strength, and tensile strain at break were obtained from the stress-strain plot. Stress was calculated by dividing the force by the initial tissue cross-sectional area, assuming a rectangular geometry (Table 1). The modulus for each sample was determined from the slope of the stress-strain curve during the apparent strain of 50–150% where the curve was almost linear while the sample had a sag during the apparent strain of 0–50%. The strain was recalculated to eliminate the sag when the Young’s modulus and the maximum strain at break.

According to the definition in the Handbook of Polymer Testing [19], “Young’s modulus” is the modulus of elasticity in tension and defined as ratio of stress difference to the corresponding strain difference (stress/strain). In this study, the initial elongation property (slope) was evaluated to determine Young’s modulus. “tensile strain at break” is defined as tensile strain at the tensile stress at break, if it breaks without yielding. “Maximum tensile stress” sustained by the test specimen during a tensile test represents tensile strength.

A-PRF/CGF/PPTF clots (1 mm thick) were compressed in the stainless-steel compressor [16] and were punched out (φ8 mm) using a biopsy punch (Kai Corp., Tokyo, Japan). After repeatedly rinsing the disks with PBS to eliminate as much serum as possible, the disks were immersed into 4 mL of 0.05% trypsin plus 0.53 mM EDTA (Invitrogen, Carlsbad, CA, USA) in a 35-mm dish inside a CO₂ incubator. Fibrin is well known to be specifically degraded by plasmin in vivo; however, because it takes a long time to determine degradation using plasmin in vitro [12] and because fibrin could be degraded also by other proteases in vivo, we used trypsin plus EDTA, which is usually used in cell culture, in this study.