Abstract
BACKGROUND: Avian influenza virus (AIV) subtype H9N2 is a low pathogenic avian influenza virus (LPAIV). OBJECTIVE: This study aims to evaluate the humoral and cellular immunity in vaccinated mice and broiler chicken by irradiated AIV antigen plus carboxymethyl chitosan bounded iron oxide nanoparticles (CMC-IO NPs) as an adjuvant. METHODS: AIV subtype H9N2 with 10(8.5) EID(50) /ml and haemagglutinin antigen assay about 10 log(2) was irradiated by 30 kGy gamma radiation dose. Then, the gamma-irradiated AIV was used as an inactivated vaccine and conjugated with CMC-IO NPs to improve immune responses on mice. IO NPs must be applied in all activated tests using 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) and N-hydroxysulfosuccinimide sodium salt (sulfo-NHS), and then functionalized by CMC as IO-CMC. Fourier transform infrared (FTIR) spectra on functionalized IO-CMC showed a peak of 638 cm(-1) which is a band between metal and O (Fe-O). RESULTS: Based on the comparison between the two X-ray diffraction (XRD) patterns on Fe(2) O(3) -NPs and IO-CMC, the characteristics of IO-NPs did not change after carboxymethylation. A CHN Analyzer was applied to measure the molecular weight of IO-CMC that was calculated as 1045 g. IO-CMC, irradiated AIV-IO-CMC and formalin AIV-IO-CMC were injected into 42 BALB/c mice in six groups. The fourth group was the negative control, and the fifth and sixth groups were inoculated by irradiated AIV-ISA70 and formalin AIV-ISA70 vaccines. An increase in haemagglutination inhibition (HI) antibody titration was observed in the irradiated AIV-IO-CMC and formalin AIV-IO-CMC groups (p < 0.05). In addition, increases in the lymphoproliferative activity of re-stimulated splenic lymphocytes, interfron-γ (IFN-γ) and interleukin-2 (IL-2) concentration in the irradiated AIV-IO-CMC group demonstrated the activation of Type 1 helper cells. The concentration of IL-4 was without any significant increases in non-group. CONCLUSIONS: Accordingly, Th2 activation represented no increase. Finally, the finding showed that AIV-IO-CMC was effective on enhancing immunogenicity as irradiated AIV antigen administered with a clinically acceptable adjuvant (i.e. IO-CMC).