Immunodominant Francisella tularensis antigens identified using proteome microarray. ^©Crown Copyright 2007 Dstl

Abstract

Stimulation of protective immune responses against intracellular pathogens is difficult to achieve using non-replicating vaccines. BALB/c mice immunized by intramuscular injection with killed Francisella tularensis (live vaccine strain) adjuvanted with preformed immune stimulating complexes admixed with CpG, were protected when systemically challenged with a highly virulent strain of F. tularensis (Schu S4). Serum from immunized mice was used to probe a whole proteome microarray in order to identify immunodominant antigens. Eleven out of the top 12 immunodominant antigens have been previously described as immunoreactive in F. tularensis. However, 31 previously unreported immunoreactive antigens were revealed using this approach. Twenty four (50%) of the ORFs on the immunodominant hit list belonged to the category of surface or membrane associated proteins compared to only 22% of the entire proteome. There were eight hypothetical protein hits and eight hits from proteins associated with different aspects of metabolism. The chip also allowed us to readily determine the IgG subclass bias, towards individual or multiple antigens, in protected and unprotected animals. These data give insight into the protective immune response and have potentially important implications for the rational design of non-living vaccines for tularemia and other intracellular pathogens.

A Burkholderia pseudomallei protein microarray reveals serodiagnostic and cross-reactive antigens

“…Our finding that these cross-reactive antigens also reacted equally with sera from individuals in the USA suggests that cross-reactive antibodies may be responsible for these signals. This finding contrasts markedly to our previous studies with Francisella tularensis (7,29), Borrelia burgdorferi (5), Plasmodium falciparum (6), and vaccinia (8), where reactivity of arrayed proteins with naïve patient sera was minimal or not seen.…”

“…We have previously developed array technology that allows protein microarrays to be constructed from the predicted proteome of a microorganism (3)(4)(5)(6)(7). These arrays can be used to address basic questions about the interactions of a given pathogen with the host immune system (8)(9)(10), and allows the identification of proteins which can be used as diagnostic reagents or for inclusion in vaccines (3).…”

Identification of Potential Serodiagnostic and Subunit Vaccine Antigens by Antibody Profiling of Toxoplasmosis Cases in Turkey

“…1B). We aimed to down-select the number of exons to ϳ2000 using a bioinformatic filtering process based on antigenic features seen in other proteome-wide serological screens in bacteria (63,64). First, genes lacking a mass spectroscopy profile were excluded to enrich for functional genes.…”

“…Meanwhile, the array employed here represents only a fraction of the nearly 8000 genes encoded in the T. gondii genome. Although the protein microarray strategy has been used extensively in bacteria (45,47,53,63,64,67,68), its application to antigen discovery in eukaryotic genomes is relatively novel, having been applied previously only to Plasmodium falciparum (46,69). Of the 15 known serodiagnostic antigens listed above, the smallest exons of four were represented on the current chip, namely GRA1 (070250_1 and _2), GRA2 (027620_2), GRA5 (086450_1), and MIC5 (077080_1).…”

Vaccinomics for the Major Blood Feeding Helminths of Humans

“…Cell-free protein expression systems, whether they be components of prokaryotic or eukaryotic cells, do not possess the cellular machinery to process secreted proteins and will not always faithfully reproduce the correct fold. Nonetheless, the high-throughput nature of this expression system lends itself well to immunomic studies and has proven to be an excellent tool for identifying antigens from a range of single celled pathogens at least (Crompton et al, 2010;Davies et al, 2008;Eyles et al, 2007).…”