In a previous study, we reported that the novel annexin XX1 (annexin E1), identical to alpha14-giardin, is specifically localized to the flagella and to the median body of the trophozoites. However, the mode of interaction and the direct partners involved remained unclear. In the present study, we show that alpha4-giardin obviously does not evenly distribute over the full length of the axonemes, but rather, resides at local slubs near the proximal part and the ends of the flagella. In immunocytochemical co-localization studies, the anti-giardin primary antibody exclusively reacted with distinct regions of the flagella in permeabilized cells, whereas the anti-tubulin antibody bound to all areas of the axonemes in the cells and to isolated cytoskeletons. Isolated cytoskeletons did not react with anti-giardin antibodies. alpha14-Giardin itself is able to assemble to multimeric structures. Taken together, our findings suggest that alpha14-giardin adheres to microtubules of the flagella via self-assembly that may regulated by Ser/Thr-phosphorylation.