Abstract
Background:
Escherichia coli bacteria are common in the environment, and the immune systems of humans and animals are essential for fighting these pathogens.

Aim:
This study aimed to produce, purify, and characterize chicken immunoglobulin Y (IgY) specific against E. coli as a passive immunity candidate.

Methods:
A single E. coli was isolated from the enteric tract of fascicularis. In this study, two groups of 22-week-old hens, each consisting of three hens, with a body weight range of 2–3 kg, were vaccinated with E. coli inactivated either with 4% formaldehyde overnight (Group A) or at 90°C for 2 hours (Group B). In the first week (priming), each chicken group was injected intramuscularly with 0.5 ml of vaccine containing E. coli at a concentration of 109 Colony Forming Unit without Montanide ISA 70 as an adjuvant. In the second week (booster), each chicken group was injected with 0.5 ml of vaccine containing E. coli at the concentration 109 CFU mixed with Montanide ISA 70 at a volume ratio of 3:7. At the third week (final booster), each chicken group was injected with 0.5 ml of vaccine containing E. coli at the concentration 109 CFU mixed with Montanide ISA 70 in a volume ratio of 3:7. Serum and egg samples were collected before booster administration at week 1 and following booster administration at weeks 2, 3, and 6.

Results:
Based on the agar gel precipitation test, the IgY titer on the chicken Group A step-by-step increase in antibody titer, with serum IgY titers of 22 and 32 at weeks 3 and 6, respectively egg yolk IgY titers of 21 and 22 at weeks 3 and 6, respectively. The IgY titer in group B had a high antibody titer, with serum IgY titers of 23 at both weeks 3 and 6, and egg yolk IgY titers of 21 and 23 at weeks 3 and 6, respectively. The results of the sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis of purified serum and egg yolk at weeks 3rd and 6th , revealed protein bands with molecular weights of approximately 65 and 25 kDa. The 65 kDa protein band is hypothesized to correspond to the heavy chain of IgY, whereas the 25 kDa band is presumed to represent the light chain of IgY. Immunoblotting of E. coli proteins with various preparations revealed approximately 29 kDa, which can be recognized by IgY, an adhesin protein.

Conclusion:
This study demonstrates that inactivated E. coli vaccination can increase IgY titers in chickens, which is a potential candidate for passive immunity against E. coli.

Key words: Chicken immunoglobulin Y; Escherichia coli; Formaldehyde inactivation; Heat inactivation.