Abstract: Self-incompatibility (SI) in Camellia oleifera Abel. represents a major biological constraint on fruit set and yield. To clarify the involvement of auxin (IAA) in SI-associated reproductive failure, two elite cultivars, ‘Huashuo’ and ‘Huajin’, were examined using integrated in vivo and in vitro approaches. Pollen tube progression following self- and cross-pollination was monitored by aniline blue staining from 24 to 96 h, endogenous IAA concentrations in pistils were determined, and responses of pollen tubes to exogenous IAA and the synthetic inhibitor 4-phenoxyphenylboronic acid (PPBo) were systematically evaluated. Results indicated that self-pollinated pollen tubes exhibited markedly reduced elongation relative to cross-pollinated counterparts, frequently arresting at the basal style region and displaying characteristic SI-related morphological abnormalities, including curling and folding. Pistils collected after self-pollination showed significantly elevated IAA levels compared with cross-pollination samples, implying that high concentrations of IAA may inhibit the growth of self-pollinated pollen tubes. In vitro pollen culture further demonstrated that high concentrations of IAA and PPBo significantly suppressed pollen germination and tube elongation, accompanied by pronounced accumulation of reactive oxygen species (ROS) and calcium ions at tube apices, reduced pectin content, disruption of actin organization, and induction of programmed cell death (PCD). Collectively, these results indicate that pollen tube development in C. oleifera requires an optimal concentration of IAA, with both excess and deficiency inhibiting growth and triggering PCD pathways. Aberrant elevation of pistil IAA following self-pollination therefore likely constitutes a key upstream signal initiating SI responses in C. oleifera.