In order to study the effect of different substrate stiffness on the attachment ability of seven-spotted ladybird beetles Coccinella septempunctata, we have performed friction experiments with female and male beetles on smooth silicone elastomer substrates of different stiffness, using a centrifugal force tester. Experiments were performed with intact beetles and those without claws. In addition, males were also tested on washed substrates to exclude effects of unpolymerized residues on the substrates. The results of the centrifuge experiment revealed that the attachment ability of females was not affected by the substrate stiffness within the range of tested stiffness. Males, however, showed a decreasing attachment ability with decreasing substrate stiffness. We argue that this sexual dimorphism may be explained by the presence of a specialized, discoidal seta type in males, which is not present in females. This type of setae, when softer than the substrate, presumably exhibits a homogeneous, peak-free interfacial stress distribution, accompanied with high pull-off forces necessary to detach these setae from the substrate. When the substrate, however, becomes softer than the discoidal setae, the interfacial stress distribution will change and exhibit stress peaks at the edges of the contact. This, in fact, will lower the pull-off forces, in agreement with the experimental results. Surprisingly, the claws had almost no influence on the attachment ability of the beetles. The present study depicts, for the first time, the effect of substrate stiffness on the attachment ability in climbing animals, whose natural habitats provide a variety of substrate stiffness ranging from rigid rock surfaces to soft, biofilm covered substrates.