title: Generation of Antigen Microarrays to Screen for Autoantibodies in Heart Failure and Heart Transplantation.
abstract: Autoantibodies directed against endogenous proteins including contractile proteins and endothelial antigens are frequently detected in patients with heart failure and after heart transplantation. There is evidence that these autoantibodies contribute to cardiac dysfunction and correlate with clinical outcomes. Currently, autoantibodies are detected in patient sera using individual ELISA assays (one for each antigen). Thus, screening for many individual autoantibodies is laborious and consumes a large amount of patient sample. To better capture the broad-scale antibody reactivities that occur in heart failure and post-transplant, we developed a custom antigen microarray technique that can simultaneously measure IgM and IgG reactivities against 64 unique antigens using just five microliters of patient serum. We first demonstrated that our antigen microarray technique displayed enhanced sensitivity to detect autoantibodies compared to the traditional ELISA method. We then piloted this technique using two sets of samples that were obtained at our institution. In the first retrospective study, we profiled pre-transplant sera from 24 heart failure patients who subsequently received heart transplants. We identified 8 antibody reactivities that were higher in patients who developed cellular rejection (2 or more episodes of grade 2R rejection in first year after transplant as defined by revised criteria from the International Society for Heart and Lung Transplantation) compared with those who did have not have rejection episodes. In a second retrospective study with 31 patients, we identified 7 IgM reactivities that were higher in heart transplant recipients who developed antibody-mediated rejection (AMR) compared with control recipients, and in time course studies, these reactivities appeared prior to overt graft dysfunction. In conclusion, we demonstrated that the autoantibody microarray technique outperforms traditional ELISAs as it uses less patient sample, has increased sensitivity, and can detect autoantibodies in a multiplex fashion. Furthermore, our results suggest that this autoantibody array technology may help to identify patients at risk of rejection following heart transplantation and identify heart transplant recipients with AMR.
authors.affiliation: Multi-Organ Transplant Program, University Health Network, Toronto, Ontario, Canada.|Division of Cardiology, University Health Network, Toronto, Ontario, Canada.
authors.forename: Andrzej
authors.initials: A
authors.lastname: Chruscinski
authors.affiliation: Multi-Organ Transplant Program, University Health Network, Toronto, Ontario, Canada.
authors.forename: Flora Y Y
authors.initials: FY
authors.lastname: Huang
authors.affiliation: Department of Immunology, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Albert
authors.initials: A
authors.lastname: Nguyen
authors.affiliation: Multi-Organ Transplant Program, University Health Network, Toronto, Ontario, Canada.
authors.forename: Jocelyn
authors.initials: J
authors.lastname: Lioe
authors.affiliation: Division of Cardiac Surgery, University Health Network, Toronto, Ontario, Canada.
authors.forename: Laura C
authors.initials: LC
authors.lastname: Tumiati
authors.affiliation: Multi-Organ Transplant Program, University Health Network, Toronto, Ontario, Canada.
authors.forename: Stella
authors.initials: S
authors.lastname: Kozuszko
authors.affiliation: Multi-Organ Transplant Program, University Health Network, Toronto, Ontario, Canada.
authors.forename: Kathryn J
authors.initials: KJ
authors.lastname: Tinckam
authors.affiliation: Multi-Organ Transplant Program, University Health Network, Toronto, Ontario, Canada.|Division of Cardiac Surgery, University Health Network, Toronto, Ontario, Canada.
authors.forename: Vivek
authors.initials: V
authors.lastname: Rao
authors.affiliation: Department of Immunology, University of Toronto, Toronto, Ontario, Canada.|Toronto General Research Institute, Toronto, Ontario, Canada.|Women's College Research Institute, Toronto, Ontario, Canada.
authors.forename: Shannon E
authors.initials: SE
authors.lastname: Dunn
authors.affiliation: Behavioral Neuroscience, Biomolecular Sciences and Human Studies Programs, Laurentian University, Sudbury, Ontario, Canada.
authors.forename: Michael A
authors.initials: MA
authors.lastname: Persinger
authors.affiliation: Multi-Organ Transplant Program, University Health Network, Toronto, Ontario, Canada.|Department of Immunology, University of Toronto, Toronto, Ontario, Canada.
authors.forename: Gary A
authors.initials: GA
authors.lastname: Levy
authors.affiliation: Multi-Organ Transplant Program, University Health Network, Toronto, Ontario, Canada.|Division of Cardiology, University Health Network, Toronto, Ontario, Canada.
authors.forename: Heather J
authors.initials: HJ
authors.lastname: Ross
mesh_terms: D000328:Adult; D000974:Antibodies, Antinuclear / Q000097:blood / Q000276:immunology; D000941:Antigens / Q000097:blood / Q000276:immunology*; D001323:Autoantibodies / Q000097:blood*; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D006084:Graft Rejection / Q000276:immunology; D006333:Heart Failure / Q000276:immunology / Q000473:pathology / Q000628:therapy*; D016027:Heart Transplantation; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood; D008297:Male; D008875:Middle Aged; D010641:Phenotype; D040081:Protein Array Analysis; D012307:Risk Factors
chemical_list: D000974:Antibodies, Antinuclear; D000941:Antigens; D001323:Autoantibodies; D007074:Immunoglobulin G