Diagnosing primary immunodeficiencies using protein-based screening assays

Abstract Primary immunodeficiencies (PIDs), now more commonly referred to as inborn errors of immunity (IEIs), are a diverse group of inherited conditions characterized by immune system defects, leading to recurrent infections, immune dysregulation, and autoimmune symptoms. Early diagnosis is crucial for effective management and treatment. This document explores the role of protein-based assays in diagnosing and characterizing IEIs. Protein-based assays, including enzyme-linked immunosorbent assay (ELISA), flow cytometry, and immunoblotting, have significantly advanced the identification and understanding of IEI. These assays enable rapid screening and functional analysis of immune proteins, providing critical insights into the phenotypic manifestations of genetic mutations. ELISA technology allows for the detection of specific antigens or antibodies, while flow cytometry facilitates the analysis of multiple cell populations and their functional states. Immunoblotting remains valuable for detecting protein expression and identifying truncated or non-functional proteins. This review highlights case studies demonstrating the application of these assays in diagnosing conditions such as X-linked lymphoproliferative syndrome type 2 (XLP2), activated phosphoinositide 3-kinase delta syndrome (APDS), Severe Combined Immunodeficiency (SCID) and Wiskott-Aldrich syndrome (WAS). It also discusses the importance of functional assays in confirming the pathogenicity of genetic variants of unknown significance (VUS) and guiding targeted genetic analysis. Overall, protein-based assays are essential tools in the diagnostic workflow for IEI, enabling timely and accurate diagnosis, guiding therapeutic decisions, and monitoring patient outcomes post-treatment, including hematopoietic stem cell transplantation (HSCT) and gene therapy.