Tensegrity structures are spatial, reticulated, and lightweight structures that are composed of struts and cables. Stability is provided by a self-stress state between tensioned and compressed members obtained by prestress. Although the form finding of tensegrity structures has attracted significant attention, few studies have focused on the structural concept implementation in large-scale systems and prestress implementation in particular. This work investigates the effects of the prestress implementation on the self-stress state in a large-scale tensegrity structure with interconnected struts. The suspended, 6.2-m-tall structure consists of stainless steel cables and carbon-fiber composite tubes. Different uniform and non-uniform prestress and support condition scenarios were applied and the effects on the self-stress state were investigated experimentally by measuring strains in the carbon tubes, stresses in cables, and reaction forces. Numerical modeling was performed using the dynamic relaxation method, whose predicted stress values were compared to experimental data. The results revealed that the cable-by-cable prestress implementation led to lower effective prestress levels than nominally predicted due to a continuous stress redistribution. Non-uniformly applied prestress resulted in a uniform stress state due to the same continuous stress redistribution. Small joint restrictions in rotational angles and small joint friction hindered geometry and stress distribution from changing when the support conditions changed. The importance of extending the analysis of tensegrity structures beyond form finding and sizing is thus demonstrated; prestress implementation and joint design also need attention.