Monoclonal antibodies to conformational epitopes of the surface glycoprotein of caprine arthritis-encephalitis virus: potential application to competitive-inhibition enzyme-linked immunosorbent assay for detecting antibodies in goat sera.
Four immunoglobulin G1 monoclonal antibodies (MAbs) to the gp135 surface envelope glycoprotein (SU) of the 79-63 isolate of caprine arthritis-encephalitis virus (CAEV), referred to as CAEV-63, were characterized and evaluated for their ability to compete with antibody from CAEV-infected goats. Three murine MAbs (MAbs GPB16A, 29A, and 74A) and one caprine MAb (MAb F7-299) were examined. All MAbs reacted in nitrocellulose dot blots with native CAEV-63 SU purified by MAb F7-299 affinity chromatography, whereas none reacted with denatured and reduced SU. All MAbs reacted in Western blots with purified CAEV-63 SU or the SU component of whole-virus lysate following denaturation in the absence of reducing agent, indicating that intramolecular disulfide bonding was essential for epitope integrity. Peptide-N-glycosidase F digestion of SU abolished the reactivities of MAbs 74A and F7-299, whereas treatment of SU with N-acetylneuraminate glycohydrolase (sialidase A) under nonreducing conditions enhanced the reactivities of all MAbs as well as polyclonal goat sera. MAbs 29A and F7-299 were cross-reactive with the SU of an independent strain of CAEV (CAEV-Co). By enzyme-linked immunosorbent assay (ELISA), the reactivities of horseradish peroxidase (HRP)-conjugated MAbs 16A and 29A with homologous CAEV-63 SU were <10% of that of HRP-conjugated MAb 74A. The reactivity of HRP-conjugated MAb 74A was blocked by sera from goats immunized with CAEV-63 SU or infected with CAEV-63. The reactivity of MAb 74A was also blocked by sera from goats infected with a CAEV-Co molecular clone, although MAb 74A did not react with CAEV-Co SU in Western blots. Thus, goats infected with either CAEV-63 or CAEV-Co make antibodies that inhibit binding of MAb 74A to CAEV-63 SU. A competitive-inhibition ELISA based on displacement of MAb 74A reactivity has potential applicability for the serologic diagnosis of CAEV infection.
Ozyörük
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Department of Veterinary Microbiology and Pathology, Washington State University, Pullman, Washington 99164-7040, USA.
Cheevers
W P
WP
Hullinger
G A
GA
McGuire
T C
TC
Hutton
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M
Knowles
D P
DP
D000581:Amidohydrolases / Q000378:metabolism; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D000914:Antibodies, Viral / Q000097:blood* / Q000276:immunology; D016088:Arthritis-Encephalitis Virus, Caprine / Q000276:immunology*; D015153:Blotting, Western / Q000379:methods; D003101:Collodion; D003429:Cross Reactions; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D018985:Epitopes, B-Lymphocyte / Q000276:immunology*; D015686:Gene Products, env / Q000276:immunology*; D006023:Glycoproteins / Q000276:immunology; D006041:Goats; D006735:Horseradish Peroxidase / Q000378:metabolism; D015151:Immunoblotting / Q000379:methods; D016180:Lentivirus Infections / Q000097:blood / Q000276:immunology / Q000821:virology*; D008565:Membrane Proteins; D051379:Mice; D008807:Mice, Inbred BALB C; D009439:Neuraminidase / Q000378:metabolism; D010084:Oxidation-Reduction; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase; D011487:Protein Conformation; D011489:Protein Denaturation; D017186:Titrimetry; D014764:Viral Proteins
D000911:Antibodies, Monoclonal; D000914:Antibodies, Viral; C412317:ENV protein, Caprine arthritis encephalitis virus; D018985:Epitopes, B-Lymphocyte; D015686:Gene Products, env; D006023:Glycoproteins; D008565:Membrane Proteins; D014764:Viral Proteins; D003101:Collodion; D006735:Horseradish Peroxidase; D009439:Neuraminidase; D000581:Amidohydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase