Healthy crabs (11.89 ± 1.2 g) were purchased from a commercial crab hatchery on Chongming Island (Shanghai, China), and transferred to the aquatic laboratory of East China Normal University and held in 120 L plastic tanks. During acclimation and toxicity bioassays, each tank was supplied with pre-aerated municipal water at 18–22 °C, pH 7.6–8.1, salinity 0.3‰, dissolved oxygen N6.5 mg/L with natural light and a photoperiod of 12 h light and 12 h dark. The crabs were fed once daily with a commercial diet (No. 9812, Shanghai Harmony Feed Co., Ltd.). Having been acclimatized for 2 weeks, 300 crabs were randomly distributed to 15 tanks. Crabs were exposed to PFOS at five nominal concentrations:0 (control), 0.01, 0.1, 1.0 and 10mg/L in a 21-day trial  while the control crabs were kept in water without adding PFOS. The range of these concentrations was based on the information derived from a preliminary experiment and represented the extreme values reported in the above mentioned studies. Perfluorooctane sulfonic acid potassium salt (CAS number 2795-39-3; N98% pure) was obtained from J&K Chemical Ltd. (Beijing, China). Each treatment included three replicates with 20 crabs each. Controls and exposures were run in a semi-static condition and the test solution was completely renewed with the corresponding concentration every day. Three crabs were sampled randomly fromeach replicate at 0, 1, 4, 7, 14 and 21 days respectively. b before sampling, crabs were anesthetized in icy water for 15 min to slow down the animal activity. The hemolymph of each individual crab was withdrawn from the base of the third pereiopod with a 1-ml sterile syringe. One part of hemolymph was diluted with an equal volume of anticoagulant (0.2 M NaCl, 0.17 M glucose, 50.00 mM trisodium citrate, 43.33 mM citric acid, 16.67 mM EDTA-Na2, pH 6.5) for hemocyte counting. The other part was treated without anticoagulant, and kept at 4 °C overnight for supernatant collection after centrifugation at 5000 × g for 10 min. Then, the hepatopancreas was removed and immediately frozen in liquid nitrogen. Serum and hepatopancreas were stored at −80 °C until further analysis.