Development of monospecific polyclonal antibodies against hypervirulent Klebsiella pneumoniae

Abstract

Darniati, Setiyaningsih S, Agungpriyono DR, Handharyani E. 2020. Development of monospecific polyclonal antibodies against hypervirulent Klebsiella pneumoniae. Biodiversitas 22: 99-105. The aim of the research was to produce monospecific antibodies against Klebsiella pneumoniae serotype K1 and K2. Klebsiella pneumonia was isolated from infected lungs of Aceh cattle. The bacteria were identified by rpoB (RNA polymerase β subunit) PCR amplification specific for K. pneumoniae. The presence of specific capsule cluster gene magA (specific to serotype K1) and K2A (specific to serotype K2) was used to characterization capsular serotype. Four New Zealand White rabbits were divided into two groups and injected with K. pneumoniae serotype K1 and K2 respectively. The presence of antibodies was determined by using AGPT and purification was performed by immunoaffinity method. The immunoreactivity was determined by direct agglutination test and immunohistochemical staining. K. pneumoniae antibodies began to appear on the 14th day after the first injection and progressively increased after the 1st and 2nd booster. The cross-reactivity between the antigen was eliminated by absorbing with the opposite antigen and several serotypes of non-K1/K2. Rabbit serum has a high protein concentration (58.485 µg/µL for K1 and 53.987 µg/µL for K2 antibody), which decreased significantly following purification (2. 377 µg / µL and 3. 721 µg / µL for K1 and K2 respectively). The SDS-PAGE characterization showed two bands with molecular weight at 54 kDa and 25 kDa representing the heavy chain and light chain. Antibodies of anti-K. pneumoniae serotypes K1 and K2 showed high affinity and specificity for homologous antigens in direct agglutination test and immunohistochemical staining.