Abstract
Canine parvovirus (CPV) can cause acute and highly contagious bloody enteritis in dog. To obtain antibodies against CPV, hens were immunized with virus-like particles (VLP) of CPV-VP2. The IgY single chain fragment variables (scFv) were generated by T7 phage display system and expressed in E. coli system. The titer of the primary scFv library reached to 1.5 × 106 pfu/mL, and 95% of the phages contained the target fragments. The CPV-VLP and CPV-VP2 protein showed similar reaction values to the purified scFv in the ELISA test, and the results of ELISA analysis using IgY-scFv toward CPV clinical samples were consistent with commercial immunochromatographic assay (ICA) and PCR detection, the scFv did not show cross reactivity with canine distemper virus (CDV) and canine coronavirus (CCV). IgY-scFv was successfully expressed in CRFK cells, and in the virus suppression assay, 55% of CPV infections were eliminated within 24 h. Docking results demonstrated that the number of amino acids of the binding sides between scFv and VP2 were AA37 and AA40, respectively. This study revealed the feasibility of a novel functional antibody fragment development strategy by generating diversified avian IgY-scFv libraries towards the pathogenic target of interest for both detection and therapeutic purposes in veterinary medicine.
Highlights
Canine parvovirus (CPV) was first identified in 1978 [1]; it has a single-stranded DNA genome of negative polarity, about 5200 bp in length
The CPV-virus-like particles (VLP) and CPV-VP2 protein showed similar reaction values to the purified single chain fragment variables (scFv) in the ELISA test, and the results of ELISA analysis using IgY-scFv toward CPV clinical samples were consistent with commercial immunochromatographic assay (ICA) and PCR detection, the scFv did not show cross reactivity with canine distemper virus (CDV) and canine coronavirus (CCV)
MAbs produced by mammals may have the side effects of immunogenicity, thrombocytopenia and hypersensitivity reactions, etc., which has greatly limited their application in target detection and treatment [14, 15]
Summary
Canine parvovirus (CPV) was first identified in 1978 [1]; it has a single-stranded DNA genome of negative polarity, about 5200 bp in length. The conventional attenuated and inactivated CPV vaccines have been successful in reducing the disease outbreaks, the genus of parvovirus still causes severe. Antibody based approach is promising in CPV disease diagnosis, therapy and prevention, and the relevant hyper immunoglobulin G (IgG) and full-length monoclonal antibody (mAbs) have been routinely applied in the veterinary practices [4]. Functional antibody fragments (i.e.: scFv, Fab) generated by phage-display technology have been not yet routinely evaluated and applied in the veterinary medicine despite it has been well confirmed that such engineered antibodies offer a series of advantages over polyclonal and full-length monoclonal antibodies. Antibody phage display technology is to display polyclonal antibodies on the surface of the phage shell protein and to screen the specific monoclonal antibodies
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