Identification of immunodominant fraction of Paramphistomum epiclitum and its evaluation for use in the serodiagnosis of paramphistomosis by ELISA

SHAFIYA IMTIAZ RAFIQI; SAKEER HUSSAIN K M; HIRA RAM; RAJAT GARG; SAROJ KUMAR; M K SINGH; P S BANERJEE

doi:10.56093/ijans.v87i12.79827

Authors

SHAFIYA IMTIAZ RAFIQI ICAR-Indian Veterinary Research Institute, Izatnagar, Uttar Pradesh 243 122 India
SAKEER HUSSAIN K M ICAR-Indian Veterinary Research Institute, Izatnagar, Uttar Pradesh 243 122 India
HIRA RAM ICAR-Indian Veterinary Research Institute, Izatnagar, Uttar Pradesh 243 122 India
RAJAT GARG ICAR-Indian Veterinary Research Institute, Izatnagar, Uttar Pradesh 243 122 India
SAROJ KUMAR ICAR-Indian Veterinary Research Institute, Izatnagar, Uttar Pradesh 243 122 India
M K SINGH ICAR-Indian Veterinary Research Institute, Izatnagar, Uttar Pradesh 243 122 India
P S BANERJEE ICAR-Indian Veterinary Research Institute, Izatnagar, Uttar Pradesh 243 122 India

https://doi.org/10.56093/ijans.v87i12.79827

Keywords:

ES antigen, Immunodiagnosis, Low molecular weight protein, Paramphistomosis

Abstract

The present study was undertaken to identify and purify the immunodominant fractions from the excretory
secretory (ES) antigen of Paramphistomum epiclitum, a predominant amphistome species infecting ruminants in
India. ES antigen was prepared and characterized using SDS-PAGE and Western blot analysis. Major polypeptides
of molecular weight 11, 22, 28, 31, 33, 39, 52, 59, 63 and 72 kDa were visualized in SDS-PAGE. Polypeptides (9)
of 11, 14, 16, 22, 31, 33, 39, 63 and 72 kDa showed immunoreactivity in Western blot analysis. The whole ES
antigen of P. epiclitum was initially concentrated using PEG-8000 followed by spin-X UF concentrator with 10
kDa cutoff range and subsequently fractionated by size exclusion chromatography using Sephadex G-25. Cross
reactivity of the P. epiclitum ES antigen was studied with positive sera of F. gigantica and H. contortus. Based on
the cross reactivity profile, the low molecular weight antigenic fraction with 11 kDa polypeptide was selected for
further use in indirect-ELISA. Bovine serum samples (258) were tested with optimized ELISA. Sensitivity of the
ELISA was calculated as 75.0%, while the specificity was 85.0%. The percent positive and negative predictive
values for the test were 70.78 and 87.57%, respectively.

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