Changes of IgM and IgG antibody levels in experimental rabbit anisakiasis as observed by ELISA and SDS-PAGE/immunoblot

, Yang, H J; , Cho, Y J; , Paik, Y H

doi:1991.29.4.389

Korean J Parasito > Volume 29(4):1991 > Article

Original Article


Korean J Parasitol. 1991 Dec;29(4):389-396. English. Published online Mar 20, 1994. http://dx.doi.org/10.3347/kjp.1991.29.4.389
Copyright © 1991 by The Korean Society for Parasitology


Changes of IgM and IgG antibody levels in experimental rabbit anisakiasis as observed by ELISA and SDS-PAGE/immunoblot
H J Yang,Y J Cho and Y H Paik
Department of Parasitology, College of Medicine, Kyung-Hee University, Seoul 130-701, Korea.


Abstract
Antibody changes in experimental anisakiasis were observed in 10 rabbits which were infected each with 10 Anisakis simplex larvae. The sera were collected before and on the 6th to the 95th day after the infection. Using crude saline extract of Anisakis larvae as antigen, specific IgM and IgG antibody levels were observed by ELISA and SDS-polyacrylamide gel electrophoresis/immunoblot. Levels of specific-IgM antibody were elevated from the 6th day, reached their peaks on the 11th day after the infection, and dropped thereafter. Serum levels of IgG antibody increased from the 6th day and reached their peak on the 26th day after the infection, and decreased gradually thereafter. When SDS-PAGE of the crude extract was done, at least forty-one SDS-polypeptide bands were recognized. Of them, IgM antibody reacted mainly to the bands of 168, 95, 74, 64, 51, 47 and 34 kDa while IgG antibody reacted strongly to 168, 92, 85, 64, 58, 52, 42 and 40 kDa bands. The crude extract showed negligible cross reactions with sera of other parasitic diseases and normal control.

Figures


	Fig. 1 Changes of the individual absorbance of Anisakis-specific IgM antibody in experimental rabbit anisakiasis.


	Fig. 2 Anisakis-specific IgG antibody levels in rabbit sera of experimental anisakiasis.


	Fig. 3 Changes of mean absorbance of Anisakis specific-IgM(+) and -IgG(▪) antibody levels (mean±S.D.) in infected rabbit sera.


	Fig. 4 SDS-PAGE/immunoblot finding of IgM reacting bands. In SDS-PAGE, 7.5~15% gradient separating gels were used. Numericals at top of the figure indicated the postinfection days. M_r: Molecular mass in kDa, C: Amidoblack B stained NC slip of the crude extract, P: Preinfection.


	Fig. 5 Finding of IgG reacting bands in SDS-PAGE/immunoblot. In SDS-PAGE, 7.5~15% gradient separating gels were used. Numericals at top of the figure indicated the postinfection days. M_r: Molecular mass in kDa, C: Amidoblack B stained NC slip of the crude extract, P: Preinfection.

Tables


	Table 1 Serum levels of Anisakis-specific IgG antibody in other parasitic infections and control

References


1.	Ashby BS, Appleton PJ, Dawson I. Eosinophilic Granuloma of Gastro-intestinal Tract Caused by Herring Parasite Eustoma Rotundatum. Br Med J 1964;1(5391):1141–1145.

2.	Desowitz RS, Raybourne RB, Ishikura H, Kliks MM. The radioallergosorbent test (RAST) for the serological diagnosis of human anisakiasis. Trans R Soc Trop Med Hyg 1985;79(2):256–259.

3.	Fujino T, et al. Jpn J Parasitol 1984;33:73–92.

4.	Hong ST, Lee SH. Histopathological and serological observations on experimental anisakiasis of rabbits. Korean J Parasitol 1987;25(2):168–180.

5.	Kim CH, Chung BS, Moon YI, Chun SH. [A case report on human infection with Anisakis sp. in Korea]. Korean J Parasitol 1971;9(1):39–43.

6.	Kim LS, Lee YH, Kim S, Park HR, Cho SY. A case of anisakiasis causing intestinal obstruction. Korean J Parasitol 1991;29(1):93–96.

7.	Kobayashi A, Tsuji M, Wilbur DL. Probable pulmonary anisakiasis accompanying pleural effusion. Am J Trop Med Hyg 1985;34(2):310–313.

8.	Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 1970;227(5259):680–685.

9.	Lee YJ, et al. Korea Univ Med J 1990;27:149–157.

*10.*	Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. Protein measurement with the Folin phenol reagent. J Biol Chem 1951;193(1):265–275.

*11.*	Oshima T. Prog Med Parasit Jpn 1972;4:305–393.

*12.*	Sakanari JA. Anisakis - from the platter to the microfuge. Parasitol Today 1990;6(10):323–327.

*13.*	Seo BS, Chai JY, Lee SH, Hong ST, Seo JW, Noh SH. A human case infected by the larva of Terranova type A in Korea. Korean J Parasitol 1984;22(2):248–252.

*14.*	Sugimachi K, Inokuchi K, Ooiwa T, Fujino T, Ishii Y. Acute gastric anisakiasis. Analysis of 178 cases. JAMA 1985;253(7):1012–1013.

*15.*	Suzuki H, et al. Jpn J parasitol 1972;21(4):252–256.

*16.*	Suzuki T, Sato Y, Yamashita T, Sekikawa H, Otsuru M. Anisakiasis: preparation of a stable antigen for indirect fluorescent antibody test. Exp Parasitol 1974;35(3):418–424.

*17.*	Suzuki T, et al. Jpn J Parasitol 1975;24(4):184–191.

*18.*	Takahashi S, Sato N, Ishikura H. Establishment of monoclonal antibodies that discriminate the antigen distribution specifically found in Anisakis larvae (type I). J Parasitol 1986;72(6):960–962.

*19.*	Tsang VC, Peralta JM, Simons AR. Enzyme-linked immunoelectrotransfer blot techniques (EITB) for studying the specificities of antigens and antibodies separated by gel electrophoresis. Methods Enzymol 1983;92:377–391.

*20.*	Bartlett A, Bidwell DE. Enzyme immunoassays for parasitic diseases. Trans R Soc Trop Med Hyg 1976;70(2):98–106.

*21.*	Yokogawa M, Yoshimura H. Clinicopathologic studies on larval anisakiasis in Japan. Am J Trop Med Hyg 1967;16(6):723–728.