Complexing amphotericin B (AmB) with lipids or entrapping it in liposomes may increase its efficacy by reducing toxicity and affecting distribution. However, depending on the lipids involved, interference with antifungal activity has been shown. We compared the in vitro activity of AmB colloidal dispersion by cholesteryl sulfate (ABCD) to the standard AmB deoxycholate suspension (ABDS) against 41 isolates of 15 pathogenic species. Broth dilution MIC and minimum fungicidal concentration (MFC) ranges were similar, and the same number of isolates had lower MICs and MFCs with one drug as with the other. Differences between species were noted. In less than a third of comparisons, there were large (fourfold or more) decreases in ABCD activity relative to that of ABDS, and in approximately 1/10, there were large increases. Thus, ABCD complexing variably affects AmB activity. As ABCD pharmacokinetics and toxicology differ from those of ABDS, the significance of these results for in vivo activity needs to be determined.