Antibody Detection of EhADH Protein

For EhADH immunodetection, we obtained hamster polyclonal antibodies (α-EhADH) against a specific EhADH peptide (N₅₆₆-QCVINLLKEFDNTKNI-C₅₈₂) localized within the adherence domain. Male hamsters were immunized three times (each 2 weeks) with 300 μg of this peptide diluted in TiterMax^® Gold Adjuvant liquid (Sigma), then, animals were bled and antibodies obtained. Other primary antibodies used were: rat α-EhVps23 (Galindo et al., 2021 (link)), mouse monoclonal α-HA (GeneTex), mouse α-EhVps32 (Avalos-Padilla et al., 2015 (link)), mouse monoclonal α-Ubiquitin (Santacruz), rabbit α-EhCP112 (García-Rivera et al., 1999 (link)) and mouse monoclonal α-human actin (kindly given by Dr. Manuel Hernández, Cell Biology Department, CINVESTAV) antibodies. Secondary antibodies were: HRP-labelled α-rabbit, α-mouse, α-hamster and α-rat IgGs (Zymed) for western blot. FITC-labelled α-rabbit and α-mouse IgGs, Alexa 647 α-hamster and TRITC-labelled or Alexa 405 α-rat IgGs (Life Technologies) for immunofluorescence. For immunoelectron microscopy experiments, we used α-rat IgG, conjugated with 10 nm gold particles (TED Pella Inc).

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Galindo A., Javier-Reyna R., García-Rivera G., Bañuelos C., Chávez-Munguía B., Salazar-Villatoro L, & Orozco E. (2022). EhVps23, an ESCRT-I Member, Is a Key Factor in Secretion, Motility, Phagocytosis and Tissue Invasion by Entamoeba histolytica. Frontiers in Cellular and Infection Microbiology, 12, 835654.

Publication 2022

TiterMax® Gold Adjuvant liquid α-rat IgG

Actin Adjuvant Animals Antibodies Fitc Gold Hamster Human Immunoelectron microscopy Immunofluorescence Male Mouse Peptide Rabbit Titermax Tritc Ubiquitin Western blot

Corresponding Organization :

Other organizations : Instituto Politécnico Nacional, Center for Research and Advanced Studies of the National Polytechnic Institute

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Variable analysis

independent variables

Immunization of male hamsters with 300 μg of the EhADH peptide (N₅₆₆-QCVINLLKEFDNTKNI-C₅₈₂) diluted in TiterMax^® Gold Adjuvant liquid (Sigma) for three times (each 2 weeks)

dependent variables

Antibodies obtained from the immunized hamsters (α-EhADH)

control variables

Positive controls: rat α-EhVps23, mouse monoclonal α-HA, mouse α-EhVps32, mouse monoclonal α-Ubiquitin, rabbit α-EhCP112, mouse monoclonal α-human actin antibodies
Negative control: Not explicitly mentioned

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