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. 2002 Oct;40(10):3771-5.
doi: 10.1128/JCM.40.10.3771-3775.2002.

Evaluation of an enzyme-linked immunosorbent assay with recombinant rhoptry-associated protein 1 antigen against Babesia bovis for the detection of specific antibodies in cattle

Suthisak Boonchit  1 , Xuenan Xuan, Naoaki Yokoyama, Will L Goff, Gale Wagner, Ikuo Igarashi
Affiliations

Evaluation of an enzyme-linked immunosorbent assay with recombinant rhoptry-associated protein 1 antigen against Babesia bovis for the detection of specific antibodies in cattle

Suthisak Boonchit et al. J Clin Microbiol. 2002 Oct.

Abstract

The gene encoding Babesia bovis rhoptry-associated protein 1 (RAP-1) was used to develop an enzyme-linked immunosorbent assay (ELISA) to measure specific antibodies against B. bovis. The B. bovis RAP-1 gene was subcloned into a baculovirus transfer vector, and the RAP-1 protein was expressed in insect cells infected with a recombinant baculovirus. The recombinant B. bovis RAP-1 of 65 kDa was detected with anti-RAP-1 mouse serum by Western blotting, and this recombinant RAP-1 was used as an antigen in the ELISA. The ELISA was able to differentiate between B. bovis-infected sera and B. bigemina-infected sera or noninfected normal bovine sera. The results demonstrate that the recombinant RAP-1 expressed in insect cells might be a useful antigen for the detection of antibodies to B. bovis.

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Figures

FIG. 1.
FIG. 1.
IFAT analysis of mouse anti-B. bovis RAP-1 antibody. Noninfected bovine erythrocytes (A) B. bovis-infected bovine erythrocytes (B), and B. bigemina-infected bovine erythrocytes (C) were reacted with mouse anti-B. bovis RAP-1 antibody.
FIG. 2.
FIG. 2.
Western blot analysis of B. bovis RAP-1 expressed in High five insect cells by using mouse anti-B. bovis RAP-1 antibody or bovine anti-B. bigemina antibody. High five insect cells (lane 1) and High five insect cells infected with AcRAP-1-Ht (lane 2) were reacted with mouse anti-B. bovis RAP-1 antibody. High five insect cells infected with AcRAP-1-Ht (lane 3) were reacted with bovine anti-B. bigemina antibody.
FIG. 3.
FIG. 3.
IFAT analysis of B. bovis RAP-1 expressed in High five insect cells by using bovine anti-B. bovis antibody or bovine anti-B. bigemina antibody. High five insect cells (A) and High five insect cells infected with AcRAP-1-Ht (B) were reacted with bovine anti-B. bovis antibody. High five insect cells (C) and High five insect cells infected with AcRAP-1-Ht (D) were reacted with bovine anti-B. bigemina antibody.
FIG. 4.
FIG. 4.
Value from ELISA with recombinant RAP-1 with experimentally infected bovine sera. Lane 1, B. bovis-infected bovine sera; lane 2, B. bigemina-infected bovine sera; lane 3, noninfected bovine sera.
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