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Fig. 2. Various algae species commonly used as larval nutrition in laboratory cultures contain thyroxine. We reared replicate samples of algae of three species [Dunaliella tertiolecta (D. tert.), Isochrysis aff. galbana (T-ISO) and Rhodomonas lens (R. lens)] at the coastal research center (WHOI) in Woods Hole (MA, USA) in summer 2002 in 25 l containers using protocols previously described by McEdward and Herrera (1999), although adapted for large-scale use. Sterile starters for all three algae species were obtained from Dr A. D. Anderson's laboratory (Woods Hole Oceanographic Institute, MA, USA). After collection of replicate samples we performed methanol:chloroform extractions and separated small molecular species using Amicon® (Bedford, MA, USA) Ultra-15 Centrifugal Filter Device (5 kDa) and then dried samples down in a Speed-Vac^TM. We re-dissolved pellets in 50 µl 0.01 mol l^–1 NaOH and measured thyroxine using ELISA (Total Thyroxine (Total T4) ELISA Kit Alpha Diagnostics, San Antonio, TX, USA) following the manufacturer's instructions. We determined total protein content for samples using the Micro BCA^TM Protein Assay Kit from Pierce (Rockford, IL, USA). Although T-ISO has an approximately 10 times smaller cell volume than D. tert. it contains approximately the same amount of thyroxine [T4 standardized by protein content (mg dl^–1 g^–1 protein)]. Differences in TH content of these algae may reflect differential effects of these algae on larval development and morphogenesis. Values are means ± 1 S.E.M. (N=3). Asterisks indicate significant difference in hormone content between samples using Student's t-test, P<0.05.

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