Boutons exhibiting vesicle mislocalization increased corresponding to the severity of jaguar loss of function. Immunostaining against synaptotagmin of NMJs on muscles 6/7 of control and jar322/Df(3R)crb87-5 third instar larvae revealed different proportions of boutons exhibiting normal and diffuse synaptotagmin staining (A and B). Images were taken at different magnifications to visualize the entire NMJ. Open arrows indicate the normal, doughnut-shaped pattern of synaptotagmin localization and closed arrows indicate the diffuse pattern of synaptotagmin localization. Scoring the synaptotagmin distribution phenotype revealed a reduction in the percentage of normal boutons and an increase in the percentage of diffuse boutons in jar loss of function mutants compared to the control (C). Number of boutons analyzed were yw n = 48, jar322/+ n = 38, Df(3R)crb87-5/+ n = 28, jar322/Df(3R)crb87-5 n = 58 and jar322/Df(3R)crb87-5 MN n = 18. Boutons used for this study were from muscle 6/7 NMJs in segments A3, A4 and A5. Bars represent mean ± SEM.