Applied biotechnology for production of immunoglobulin Y specific to hepatitis A virus.
A new protocol for producing polyclonal antibody against hepatitis A virus (HAV) is described. Twenty hens were immunized three times with a commercial HAV vaccine and HAV from a cell culture with three types of adjuvants: CpG oligodeoxynucleotides (CpG-ODN), incomplete Freund's adjuvant and an alum adjuvant. In each of the last two booster inoculations, blood from the birds was collected and tested for HAV antibodies. Egg yolk was separated from egg white and immunoglobulin Y (IgY) antibody was then purified by polyethylene glycol 6000. The mean yield of total protein in yolk was 22.62 mg/mL. Specific activity of the antibody was tested using commercial ELISA, Western blotting, and in vitro neutralization assay demonstrating that anti-HAV IgY bound specifically. After the first immunization, birds immunized with HAV from cell culture plus incomplete Freund's adjuvant with/without CpG-ODN showed highest levels of anti-HAV IgY in serum (p<0.05). Viral combination with CpG-ODN resulted in early response of anti-HAV serum in hens, reflecting the amount of IgY transferred to the egg yolk (p<0.05). The results suggest that egg yolk may be a large scale source of specific antibodies against hepatitis A virus. Further applications of this method have yet to be tested.
de Paula
Vanessa Salete
VS
Oswaldo Cruz Institute/FIOCRUZ, Rio de Janeiro, RJ, Brazil.
da Silva
Alexandre dos Santos
Ados S
de Vasconcelos
Gentil Arthur Lins Bentes Mendonça
GA
Iff
Ezio Tavares
ET
Silva
Maria Eduarda Monteiro
ME
Kappel
Livia Abud
LA
Cruz
Paula Borba
PB
Pinto
Marcelo Alves
MA
D000276:Adjuvants, Immunologic / Q000008:administration & dosage; D000818:Animals; D001709:Biotechnology / Q000379:methods*; D015153:Blotting, Western; D002645:Chickens; D004797:Enzyme-Linked Immunosorbent Assay; D035922:Hepatitis A Antibodies / Q000276:immunology / Q000302:isolation & purification*; D022362:Hepatitis A Vaccines / Q000008:administration & dosage / Q000276:immunology; D030041:Hepatitis A virus / Q000276:immunology*; D007117:Immunization, Secondary / Q000379:methods; D007136:Immunoglobulins / Q000276:immunology / Q000302:isolation & purification*; D010063:Ovum / Q000276:immunology; D014611:Vaccination / Q000379:methods
D000276:Adjuvants, Immunologic; D035922:Hepatitis A Antibodies; D022362:Hepatitis A Vaccines; C016512:IgY; D007136:Immunoglobulins