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Korean J Parasitol > Volume 24(2):1986 > Article

Original Article
Korean J Parasitol. 1986 Dec;24(2):187-193. English.
Published online Mar 20, 1994.  http://dx.doi.org/10.3347/kjp.1986.24.2.187
Copyright © 1986 by The Korean Society for Parasitology
ELISA of paragonimiasis in cat by crude and purified antigens of Paragonimus westermani
Ok Ran Lee and Jae Kyung Chang
Department of Parasitology, Soonchunhyang University College of Medicine, Korea.
Department of Parasitology, College of Medicine, Yonsei University, Korea.
Abstract

Enzyme-linked immunosorbent assay (ELISA) using crude and affinity-purified antigens of adult worms of Paragonimus westermani was performed for infected cat sera with different worm burden, from preinfection to 18th week after infection. Crude antigen was used with supernatant of homogenated worms by freezing-thawing method, and the supernate was centrifuged for l hour at 10,000 rpm at 4C. Affinity-purified antigen (antibody-bound antigen) was prepared from fractions (bound and unbound) of crude antigen by affinity chromatography on CNBr-activated sepharose 4B, and IgG as a ligand was prepared from paragonimiasis cat serum (6 months infected) obtained by ammonium sulfate (40-45 per cent saturated) precipitation method. By SDS-PAGE, crude antigen showed 22 polypeptide fractions while purified antigen showed 4 fractions: 36, 400, 34,700,27,600 and 11,500 in molecular weights. All cats were divided into five groups(G1-G5) by different worm burdens. The mean of recovered worms (±SD) and the number of cats in each group are as follows:G1, 2 worms(0) and 4 cats; G2, 4.75(±0.66) and eight; G3, 10.75(±1.92) and four; G4, 25.20(±3.43) and five; G5, 48(±12.63) and five cats. The results were summarized as follows: The antibody levels(OD value) increased by worm burden in G1 to G4 generally. However, individual antibody levels were not exactly related with worm burden in all groups, especially there was a wide difference in G4 and G5. These results suggested that the worm burden in G4 (about 20 - 30 worms) is enough to produce antibody maximum in cats of 2~3 kg weight. The antibody levels increased significantly (p<0.05) compared to control sera at the 3rd week in G1 and G2, at the 2nd week in G3, and at the 1st week in G4 and G5. Especially in the 4th week, OD value increased more in G1(p<0.001) and in G2 to G5(p<0.01). In the pattern of antibody levels by ELISA in each group, OD in G1 increased to the 18th week continuously, in G2 OD was maintained same after the 16th week, but in G3 it decresed after the 16th week, and it was maintained same in G4 and G5 after the 14th week. The antibody levels by ELISA with the affinity-purified antigen were higher than those with crude antigen in all groups generally. Especially, the difference of OD values between two antigens was larger from the 4th to the 10th week. In G1 and G2 OD with purified antigen was higher than that with crude one to the 18th week. It was also higher in G3 than that with crude antigen to the 16th week and OD of G4 and G5 were higher before the 14th week than that with crude antigen, however became lower at the 16th week. Consequently, the antibody level in ELISA with affinity-purified antigen was more sensitive at the early weeks after infection and in light infection groups than that with crude antigen.

Figures


Fig. 1
Affinity chromatography of crude worm extract of Paragonimus westermani using CNBr-activated Sepharose 4B.


Fig. 2
Electrophoretic analysis of crude and affinity-purified antigens of Paragonimus westermani


Fig. 3
Antibodies as detected by crude antigen of P. westermani; the specific IgG antibody titers in sequence from cats experimentally infected with worms of 2(G1), 4~6(G2), 9~14(G3), 18~28(G4), 34~69(G5) were measured suing ELISA.


Fig. 4
IgG antibody titer curves in ELISA of paragonimiasis westermani cat sera against crude and purified antigens of P. westermani

Tables


Table 1
Numbers of recovered worms in each group of experimental cat paragonimiasis

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