Infectious diseases represent a major cause of morbidity and mortality worldwide. Early detection of infections is capital for managing life-threatening cases. So far, traditional diagnostic methods such as microbiological cultures are slow and, sometimes, inaccurate. In the molecular era, high-throughput techniques are essential for providing tools that are able to diagnose in a fast and reliable way, as well as they can be used for monitoring the humoral response of groups of people in a program of epidemiological surveillance when an outbreak occurs, or when a vaccine is being evaluated. Antigen-based protein microarrays are an ideal means for these purposes, as they can carry up to thousands of protein antigens from pathogenic sources and be probed with sera from different human groups (acute or chronic infected people, convalescent, controls). For the diagnosis of bacterial infections, the best antigens are in principle the surface proteins, as they have the highest chances to raise an effective immune response. Here we describe a general protocol for fabricating a glass slide-based protein microarray using recombinant bacterial surface antigens, according to our own expertise in the study of pneumococcal disease. The probing with human sera aims to evaluate differences between diseased and healthy people, in order to discover discriminating antigens that can be used, after appropriate validation, in further easy-to-use formats such as immunostrips.
Keywords: Antibodies; Diagnostics; Humoral response; Infectious diseases; Protein array; Serological assay.