Characterization of monoclonal antibody specific to entamoeba histolytica cells and erythrophagosomes
3
Issued Date
2014
Resource Type
Language
eng
Rights
Mahidol University
Suggested Citation
Tanpitcha Phongchaipaiboon, Waroon Boonyaudomsart, Saengduen Moonsom, แสงเดือน มูลสม (2014). Characterization of monoclonal antibody specific to entamoeba histolytica cells and erythrophagosomes. Retrieved from: https://repository.li.mahidol.ac.th/handle/123456789/63144
Title
Characterization of monoclonal antibody specific to entamoeba histolytica cells and erythrophagosomes
Other Contributor(s)
Abstract
Objective: Entamoeba histolytica is a pathogenic protozoan which causes
the third most deaths from parasitic diseases worldwide. Monoclonal
antibodies (mAbs), which are specific to E. histotytica were successfully
produced previously. In this study, E. histolytica specific mAbs were characterized for
localization of mAb-binding partner in the live E. histolytica trophozoites.
Methods: Trophozoite cells were stained with a mAb and analyzed by fluorescent
microscopy and flow cytometry, to investigate the localization of a mAb reacting
protein in the pathogen cells. mAbs were further tested with proteins extracted from
cellular fractions of E. histolytica using ELISA. Trophozoites were optimally challenged
with red blood cells and proteins were solubilized from the erythrophagocytosed cells
by detergents. Soluble proteins were incubated with protein G-beads linked mAb#32
or a control mAb#19. E. histolytica proteins pulled down by mAbs were analyzed with
SDS-PAGE and visualized by silver stain.
Results and Discussion: The results showed that most of the mAbs bind to cytoplasmic
proteins, with 4 clones interacting with the membrane proteins and two clones reacting
with phagosomes located inside the cells. It was found that challenging E. histolytica
cells with 0.25% human red blood cells for 1 hour at 37°C resulted in the highest yield of
the erythrophagosomes. SDS-PAGE revealed E. histolytica proteins with a molecular
masses of about 60-65 kDa and more than 170 kDa, which are successfully pulled
down by mAb #32. These proteins will be subjected to further protein identification by
mass spectrometry and may play role erythrophagocytosis, which is a virulent factor
of the parasite during tissue invasion.
Description
Joint International Tropical Medicine Meeting 2014: 3D perspectives on tropical medicine: drivers, diversity and determination the 8th seminar on food-and water-borne parasitic zoonoses: 2-4 December 2014: Centara Grand Bangkok Convention Center at Central World, Bangkok, Thailand. Bangkok: Faculty of Tropical Medicine, Mahidol University; 2014. p. 237.