Plasma volume (PV), determined indirectly from changes in haematocrit (Hct) and haemoglobin concentration ([Hb]), underestimates the absolute PV change (Evans blue dye) during thermoneutral immersion. Since PV changes during cold-water immersion have only been determined indirectly, we hypothesised that a similar underestimation may occur. Therefore, we compared the indirectly-measured PV with a direct-tracer dilution method (Evans blue dye column elution) in seven healthy males, during three, 60-min exposures: air (control; 21.2 degrees C), thermoneutral immersion (34.5 degrees C) and cold-water immersion (18.6 degrees C). During thermoneutral immersion, the directly-measured PV increased by 16.2 (1.4)% (P<0.05) and the indirectly-measured by 8.5 (0.8)% (P<0.05), with the latter underestimating the former by 43 (9.1)% (P<0.05). During cold immersion, the direct PV decreased by 17.9 (3.0)% (P<0.05) and the indirect by 8.0 (1.2)% (P<0.05), with the latter representing a 52 (6.8)% (P<0.05) underestimation of the direct PV change. Directionally-equivalent underestimations of PV change occur when using the indirect method during both thermoneutral and cold-water immersion. The assumptions inherent in the indirect method (constant F-cell ratio) appear to be violated during water immersion.