Purification of vitellin from the ovary of Chinese mitten-handed crab (Eriocheir sinensis) and development of an antivitellin ELISA

Chen, Liqiao; Jiang, Hongbo; Zhou, Zhiguang; Li, Kang; Li, Kai; Deng, Glenn Y; Liu, Zhanjiang

doi:10.1016/j.cbpc.2004.04.012

Cited by 32 publications

(21 citation statements)

References 34 publications

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“…However, depending on species or the procedure of extraction, vitellin may disintegrate into subunits. Serrano-Pinto et al (2003) found subunits in the range of 75-180 kDa in crayfish and Chen et al (2004) found two subunits of 97 and 74 kDa in Chinese mitten-handed crabs after denaturing SDS-PAGE. In our study we did not identify vitellin immunologically in eggs and larvae of L. santolla, but we assume that larvae catabolize endogeneous vitellin like yolk proteins which are similar to the yolk proteins from the eggs.…”

Section: Discussionmentioning

confidence: 99%

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Digestive enzymes in the ontogenetic stages of the southern king crab, Lithodes santolla

et al. 2006

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The early ontogenetic stages of the sub-Antarctic king crab Lithodes santolla were analyzed for the presence and activities of a set of important digestive enzymes. The eggs and non-feeding larvae (zoea I-III, megalopa) showed high activities of esterases, phosphatases, and exopeptidases indicating the enzymatic ability to utilize endogeneous yolk reserves. SDS-PAGE showed a continuous decrease of proteins or proteids in the range of 59-81 kDa during ontogenetic development from the eggs through the zoeal stages to the first juvenile crab stage, CI. This reduction reflects the degradation of storage compounds during lecithotrophic larval development. Activities of the endopeptidases, trypsin and chymotrypsin, were low in eggs and larvae but increased significantly in the first juvenile crab stage. These enzymes typically facilitate the first steps of proteolysis in the extra-cellular spaces of the midgut gland and in the stomach. Their scarcity indicates that the larvae of L. santolla are physiologically not prepared to digest external food. This ability seems to appear first in the CI stage. Extracts of juvenile midgut glands and the gastric fluids of adults showed high activities of a variety of digestive enzymes including phosphatases, carbohydrases, as well as endo-and exopeptidases. High activities of digestive enzymes in adults may compensate for scarce food supply and rate-limiting low temperatures in the predominantly sub-Antarctic habitats of L. santolla.

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Section: Discussionmentioning

confidence: 99%

“…Vitellin, which is a major yolk component in crustacean eggs, shows native molecular masses between 300 and 600 kDa (e.g. Tom et al 1992;Serrano-Pinto et al 2003;Chen et al 2004). However, depending on species or the procedure of extraction, vitellin may disintegrate into subunits.…”

Section: Discussionmentioning

confidence: 99%

Digestive enzymes in the ontogenetic stages of the southern king crab, Lithodes santolla

et al. 2006

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“…plumulosus . In the Chinese mitten-handed crab, Eriocheir sinensis , Chen et al [25] reported a similar working range of 8–500 ng/ml using the VTN polyclonal antibody-based ELISA. Ghekiere et al [28] showed that the working range of mysid VTN ELISA was 4–500 ng/ml using VTN polyclonal antibody.…”

Section: Discussionmentioning

confidence: 99%

“…The analysis of direct coating ELISA was carried out according to the method of Chen et al [25]. Polystyrene microtiter plate (Corning, USA) was coated with purified MT.…”

Section: Methodsmentioning

confidence: 99%

Expression, purification of metallothionein genes from freshwater crab (Sinopotamon yangtsekiense) and development of an anti-metallothionein ELISA

et al. 2017

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Using the phoA-fusion technology, the recombinant metallothionein (MT) from freshwater crab (Sinopotamon yangtsekiense) has been successfully produced in Escherichia coli. MT purified from the bacterial suspension showed one polypeptide with a molecular weight of 7 kDa by tricine-sodium dodecyl sulfate-polyacrylamide gel electrophoresis (Tricine-SDS-PAGE). Western-blotting confirmed the polypeptides had a specific reactivity with mouse polyclonal MT anti-serum. Based on the purified MT and MT anti-serum, the reaction parameters for an enzyme-linked immunosorbent assay (ELISA) were developed. The direct coating ELISA showed a higher linear relationship compared to antibody sandwich coating ELISA. The optimal dilution rates of purified MT anti-serum and coating period were shown to be 1:160,000 and 12 hours at 4°C. At 37°C, the appropriate reaction duration of the first antibody and the second antibody were 2 hours and 1 hour, respectively. According to these optimal parameters, the standard linear equation, y = 0.0032x + 0.1769 (R2 = 0.9779, x, y representing MT concentration and OD450 value), was established for the determination of MT concentration with a valid range of 3.9–500 ng/ml. In verification experiments, the mean coefficients of variation of the intra-assay and inter-assay were 3.260% and 3.736%, respectively. According to the result of MT recovery, ELISA with an approaching 100% MT recovery was more reliable and sensitive than the Cd saturation assay. In conclusion, the newly developed ELISA of this study was precise, stable and repeatable, and could be used as a biomarker tool to monitor pollution by heavy metals.

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“…The main techniques for VTG determination currently are enzyme linked immunosorbent assay (ELISA) [7][8][9], radioimmunoassay (RIA) [10,11], Western blots [12], HPLC [13] and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) [14]. In many related articles, immunoassays were applied in VTG analysis as the commonly used approach.…”

Section: Introductionmentioning

confidence: 99%

Separation and detection of vitellogenin in fish plasma by capillary zone electrophoresis

Song

Shao

et al. 2005

Journal of Chromatography B

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Purification of vitellin from the ovary of Chinese mitten-handed crab (Eriocheir sinensis) and development of an antivitellin ELISA

Cited by 32 publications

References 34 publications

Digestive enzymes in the ontogenetic stages of the southern king crab, Lithodes santolla

Digestive enzymes in the ontogenetic stages of the southern king crab, Lithodes santolla

Expression, purification of metallothionein genes from freshwater crab (Sinopotamon yangtsekiense) and development of an anti-metallothionein ELISA

Separation and detection of vitellogenin in fish plasma by capillary zone electrophoresis

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