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Analysis of protein components at varioue stages of Clonorchis sinensis
Won-Young Choi,Young-Kwan Jin,Ok-Ran Lee and Woon-Gyu Kim
Department of Parasitology, Catholic Medical College, Seoul, Korea.
Abstract
In this study the authors examined the protein components at various stages of Clonorchis sinensis, and those of tegument and metabolite of adult Clonorchis by using SDS-polyacrylamide gel electrophoresis and immunodiffusion. The following results were obtained:
1. The protein components of C. sinensis were gradually changed during its development. A considerable change occurred during the initial 7 days after the metacercarial infection.
2. Two bands of protein of about 97,000 molecular weight (MW) and 178,000 MW were unique to excysted metacercaria of C. sinensis. Other 2 bands of protein of 23,000 and 25,000 MW which were absent in metacercariae, might be associated with the development of sex organs in adult.
3. In the metabolite, some components of tegumental proteins were detected. And this tegumental protein components in metabolite seems to be the major antigenic components reacting with infected rabbit antiserum by immunodiffusion.
4. Twenty bands of protein were detected in the isolated adult tegument. Among them 6 bands were in 97,000~65,000 MW, 3 bands in 56,000~53,000 MW and 5 bands in 37,000~30,000 MW. On the other hand, in metabolic products of adult C. sinensis, 17 bands were detected.
Figures
Fig. 1 Electrophoretic mobility of standard proteins as a function of molecular weight (10% acrylamide).
The mobility relative to the dye front is shown for the standard proteins of Fig. 4. See explanation of Fig. 4 for molecular weight values.
Fig. 2 Precipitin reacton of infected rabbit antiserum with clonorchis antigens in agar gel.
Center well: Positive control serum (PCS)
Outer wells: 1; P1 fraction of metacercaria 2; S1 fraction of metacercaria 3; S1 fraction of 18-days CS 4; S1 fraction of 22-days CS 5; S1 fraction of 30-days CS 6; S1 fraction of 90-days CS
Fig. 3 Precipitin reaction of infected rabbit antiserum with Clonorchis antigens in agar gel.
Center well: Positive Control Serum (PCS)
Outer wells: 1; S1 fraction of 90-days CS 2; P3 fraction of 90-days CS 3; S3 fraction of 90-days CS 4; MP1 (Metabolic Products in saline) 5; MP2 (Metabolic Products in Tyrode's solution) 6; SM (Surface Membrane)
Fig. 4 Electrophoretic gel pattern of Clonorchis sinensis of various developmental stages in 10% acrylamide.
The samples, from left to right, contained 1500, excysted metacercaria(mc); 140, 7-days CS; 100, 10-days CS; 50, 14-days CS; 15, 18-days CS; 10, 22-days CS;8, 30-days CS; 2, 90-days CS; and soluble protein fraction (S3) of 90-days CS.
Standard proteins (SP), from top to bottom, phosphorylase b (94,000MW), bovine serum albumin (67,000MW), ovalbumin (43,000MW), carbonic anhydrase (30,100MW), soybean trypsin inhibitor (20,000MW) and α-lactalbumin (14,400MW).
Fig. 5 Densitometric recordings of electrophoretic gel pattern of Clonorchis sinensis of various developmental stages.
Fig. 6 electrophoretic gel pattern of surface membrane (SM), metabolic product (MP1, MP2) and 90-days worms (90) Clonorchis sinensis (10% acrylamide). Polypeptide bands were numbered according to an arbitrary scheme and their apparent molecular weights estimated by comparison with standard proteins.
Tables
Table 1 Molecular weights of the electrophoretic bands, indicated in Fig. 5, which change significantly their relative protein amounts during development of Clonorchis sinensis.
Table 2 Molecular weights of the electrophoretic bands of surface membrane and metabolic products of Clonorchis sinensis, indicated in Fig. 6
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