Hyphae ofSaprolegnia ferax continue growing at up to normal rates after staining with the fluorescent Ca2+ indicator, chlortetracycline (CTC). These hyphae show tip-high gradients of fluorescence comparable to those of other tip-growing cells. The dominant, intensely stained structures, are mitochondria. Thus, the fluorescence gradients primarily indicate the abundance of mitochondria. Sub-apically, transvacuolar strands and nuclei stain with low intensity, suggesting that tonoplasts and endoplasmic reticulum (nuclear envelope) are associated with free Ca2+ but that mitochondria dominate Ca2+ distribution within hyphae. Mitochondrial Ca2+ content is responsive to exogenous Ca2+ concentration. A previously unreported feature of the hyphal tip, which differs from observations on some pollen tubes, is the absence of fluorescence in the extreme tip where growth and wall vesicle membrane concentration are maximal and growth associated Ca2+ influxes have been postulated. CTC also shows time-dependent alteration of mitochondrial morphology which correlates with some reduction in average growth rate, even though many hyphae continue to grow at normal rates. These observations suggest that (a) mitochondria, not vacuoles, are dominant in Ca2+ sequestration in the hyphae; (b) there is heterogeneity in Ca2+-related behavior among tip growing cells; and (c) mitochondrial morphology is not critical to normal tip growth. We also discuss a number of seldom considered limitations on the use of CTC as a Ca2+ indicator. © 1991 Academic Press, Inc.