Fig. 3. (A-D) Effect of cell membrane disrupters on maximally stiffened samples (i.e. protocol 1). (A) Effect of immersion for 2.5-3 h in ASW (control; CON), 1% Triton X-100 (TRI), 0.1% Quillaja saponin (SAP), deionised water (DW) and freezing at -24°C for 3 days followed by thawing (FRO). In all cases where statistics could be applied (i.e. where the standard deviation was not zero), P<0.01. (B) Reversibility of effects of membrane disrupters. After treatment and testing, all samples used for the experiments in A were left in ASW for 3.5 h then retested. In all cases where statistics could be applied, P<0.01. (C) Effect of immersion for 2 h in 0.05% nystatin (NYS). (D) Reversibility of effect of nystatin. After treatment and testing, the samples used in the experiments in C were left in ASW for 2 h then retested. (E) Effect of membrane disrupters on partially destiffened samples (i.e. protocol 2). After excision, samples were left for 5-6 h in ASW then tested (solid bars). They were then left for 2 h in the stated media (CON=ASW) and retested (cross-hatched bars). In A-D, statistical comparison is between mean deflections of ASW control and treated samples; in E, statistical comparison is between mean deflections of each group of samples before (solid bars) and after (cross-hatched bars) treatment. Vertical bars represent standard deviations and asterisks indicate statistically significant differences between means: ^*P<0.05, ^**P<0.01, ^***P<0.001.