Multiplex Serological Assay for T. cruzi

Macaque plasma reactivities against 9 recombinant T. cruzi proteins and parasite lysate were tested in a multiplex assay previously described for testing human and dog serum samples [20 (link),22 (link)]. Briefly, macaque plasma was diluted 1:500 and incubated with a pool of the recombinant T. cruzi proteins attached to addressable Liquichip Ni-NTA beads (Qiagen, CA) and T. cruzi amastigote/trypomastigote lysate coupled to Carboxy-Ni-NTA beads (Qiagen Inc). Antibody binding was detected with goat anti-human IgG conjugated to phycoerythrin and quantified on a Bio-Plex Suspension Array System (Bio-Rad, Hercules, CA, USA). Weighted median fluorescence intensity (MFI) for samples in duplicate was calculated and the ratio of the specific MFI for each antigen versus a negative control (green fluorescent protein) was estimated for each antigen.

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Padilla A.M., Yao P.Y., Landry T.J., Cooley G.M., Mahaney S.M., Ribeiro I., VandeBerg J.L, & Tarleton R.L. (2021). High variation in immune responses and parasite phenotypes in naturally acquired Trypanosoma cruzi infection in a captive non-human primate breeding colony in Texas, USA. PLoS Neglected Tropical Diseases, 15(3), e0009141.

Publication 2021

Liquichip Ni-NTA beads Carboxy-Ni-NTA beads Bio-Plex Suspension Array System

Anti igg Antibody Antigen Assay Fluorescence Goat Green fluorescent protein Human Macaque Parasite Phycoerythrin Plasma Plasma proteins Recombinant proteins Serum T cruzi

Corresponding Organization : Drugs for Neglected Diseases Initiative

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Variable analysis

independent variables

Macaque plasma diluted 1:500

dependent variables

Antibody binding to recombinant T. cruzi proteins and T. cruzi amastigote/trypomastigote lysate, detected with goat anti-human IgG conjugated to phycoerythrin
Weighted median fluorescence intensity (MFI) for samples in duplicate
Ratio of the specific MFI for each antigen versus a negative control (green fluorescent protein)

control variables

Recombinant T. cruzi proteins attached to addressable Liquichip Ni-NTA beads
T. cruzi amastigote/trypomastigote lysate coupled to Carboxy-Ni-NTA beads

controls

Negative control: Green fluorescent protein

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