TY - JOUR
T1 - Shotgun Characterization of the Circulating IgM Antigenome of an Infectious Pathogen by Immunocapture-LC-MS/MS from Dried Serum Spots
AU - Abad, Paloma
AU - Coronado, Montserrat
AU - Vincelle-Nieto, África
AU - Pérez-Benavente, Susana
AU - Fobil, Julius N.
AU - Puyet, Antonio
AU - Diez, Amalia
AU - Reyes-Palomares, Armando
AU - Azcárate, Isabel G.
AU - Bautista, José M.
N1 - Publisher Copyright:
© 2024 American Chemical Society.
PY - 2024/2/2
Y1 - 2024/2/2
N2 - One of the main challenges in compiling the complete collection of protein antigens from pathogens for the selection of vaccine candidates or intervention targets is to acquire a broad enough representation of them to be recognized by the highly diversified immunoglobulin repertoire in human populations. Dried serum spot sampling (DSS) retains a large repertoire of circulating immunoglobulins from each individual that can be representative of a population, according to the sample size. In this work, shotgun proteomics of an infectious pathogen based on DSS sampling coupled with IgM immunoprecipitation, liquid chromatography-mass spectrometry (LC-MS/MS), and bioinformatic analyses was combined to characterize the circulating IgM antigenome. Serum samples from a malaria endemic region at different clinical statuses were studied to optimize IgM binding efficiency and antibody leaching by varying serum/immunomagnetic bead ratios and elution conditions. The method was validated using Plasmodium falciparum extracts identifying 110 of its IgM-reactive antigens while minimizing the presence of human proteins and antibodies. Furthermore, the IgM antigen recognition profile differentiated between malaria-infected and noninfected individuals at the time of sampling. We conclude that a shotgun proteomics approach offers advantages in providing a high-throughput, reliable, and clean way to identify IgM-recognized antigens from trace amounts of serum. The mass spectrometry raw data and metadata have been deposited with ProteomeXchange via MassIVE with the PXD identifier PXD043800.
AB - One of the main challenges in compiling the complete collection of protein antigens from pathogens for the selection of vaccine candidates or intervention targets is to acquire a broad enough representation of them to be recognized by the highly diversified immunoglobulin repertoire in human populations. Dried serum spot sampling (DSS) retains a large repertoire of circulating immunoglobulins from each individual that can be representative of a population, according to the sample size. In this work, shotgun proteomics of an infectious pathogen based on DSS sampling coupled with IgM immunoprecipitation, liquid chromatography-mass spectrometry (LC-MS/MS), and bioinformatic analyses was combined to characterize the circulating IgM antigenome. Serum samples from a malaria endemic region at different clinical statuses were studied to optimize IgM binding efficiency and antibody leaching by varying serum/immunomagnetic bead ratios and elution conditions. The method was validated using Plasmodium falciparum extracts identifying 110 of its IgM-reactive antigens while minimizing the presence of human proteins and antibodies. Furthermore, the IgM antigen recognition profile differentiated between malaria-infected and noninfected individuals at the time of sampling. We conclude that a shotgun proteomics approach offers advantages in providing a high-throughput, reliable, and clean way to identify IgM-recognized antigens from trace amounts of serum. The mass spectrometry raw data and metadata have been deposited with ProteomeXchange via MassIVE with the PXD identifier PXD043800.
KW - antigens
KW - immunoglobulin
KW - immunomics
KW - immunoprecipitation
KW - infection
KW - malaria
UR - http://www.scopus.com/inward/record.url?scp=85182001766&partnerID=8YFLogxK
U2 - 10.1021/acs.jproteome.3c00439
DO - 10.1021/acs.jproteome.3c00439
M3 - Article
C2 - 38183416
AN - SCOPUS:85182001766
SN - 1535-3893
VL - 23
SP - 633
EP - 643
JO - Journal of Proteome Research
JF - Journal of Proteome Research
IS - 2
ER -