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Haemagglutination inhibition (HI) assay

AS Angita Shrestha
RM Rick Meeuws
JS Jean-Remy Sadeyen
PC Pengxiang Chang
MH Marielle Van Hulten
MI Munir Iqbal
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Whole blood was collected from the vaccinated chickens and the sera separated by centrifugation at 1500 × g for 30 min at 4 °C. The HI assay was performed in V-bottomed microtiter plates following World Health Organisation guidelines61. Briefly, two-fold serial dilution of the serum was prepared by mixing 25 μl of serum with 25 μl phosphate-buffered saline (PBS). Next, 4 HAU (in 25 μl) of UDL 01/08 H9N2 virus was added to the diluted serum and incubated at 37 °C for 1 h. Finally, 50 μl of 1% chicken red blood cells were added onto the serum-virus mixture and incubated at room temperature for 45 min. The HI titres were expressed as reciprocal of the highest dilution of serum that causes total inhibition of 4 HAU of virus haemagglutination activity.

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