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Sheet 2. is currently able to detect infections within the last 9-months with 80% sensitivity and 80% specificity. Further work is required to optimize protein expression and protein constructs used for antibody detection. Methods The antibody response against the top performing predictor of recent infection, reticulocyte binding protein 2b (PvRBP2b), was tested against multiple fragments of different sizes and from different expression systems. The IgG induced against the recombinant PvRBP2b fragments in infected individuals was measured at the time of infection and in a year-long observational cohort; both conducted in Thailand. Results The antibody responses to some but not all different sized fragments of PvRBP2b protein are highly correlated with each other, significantly higher 1-week post-infection, and show potential for use as predictors of recent infection. Conclusions To achieve elimination goals, novel diagnostics are required to aid in detection of hidden parasite reservoirs. PvRBP2b was previously shown to be the top candidate for single-antigen classification of recent exposure and here, it is concluded that several alternative recombinant PvRBP2b fragments can achieve equal sensitivity and specificity at predicting recent exposure. Supplementary Information The online version contains supplementary material available at 10.1186/s12936-022-04085-x. is the worlds most widely distributed species of to infect humans causing malaria, and in near- and pre-elimination settings is proving more challenging to successfully eliminate [1]. In 2019, there were an estimated 229?million cases of malaria across all five of the World Health Organization (WHO) regions [2]. Cases of both and are seen in all regions, however infection from is overwhelmingly the predominant cause of malaria (93% of all cases), driven particularly by infections throughout sub-Saharan Africa [2]. Despite this, as Isoconazole nitrate total malaria cases decrease, the proportion of infections that are attributable to outside Africa is increasing and overall reductions in case rates are slower for is a relapsing form of matures towards the infective blood-stage by forming hepatic schizonts, but can also become developmentally arrested as a hypnozoite making an individual at risk of future relapsing episodes either weeks or months after the initial infection (when an individual is not treated with anti-hypnozoite drugs) [5, 6]. Most blood-stage infections, which also produce transmissible gametocytes, have been shown to be the product of a relapsing hypnozoite as opposed to new transmission events of primary infection [7C9]. Current diagnostics to detect infection in programmatic settings rely on either the use of rapid diagnostic tests (detecting parasite antigens in the blood) or microscopy of Giemsa-stained blood-smears [2]. Both these methods will only detect a current blood-stage infection, hence missing all asymptomatic hypnozoite infections. These methods are also insensitive to the common low parasitaemic infections of elimination, the overwhelming majority of infectious episodes that are caused by relapsing hypnozoite infections reiterates the need to innovatively target this reservoir of disease [11]. The ability to identify individuals who are infected with hypnozoites would be revolutionary in control, however the sparsity of infection throughout the liver and the inaccessibility of this organ makes this exceptionally challenging [12, 13]. Previous work has identified a novel panel of eight recombinant antigens that can be used to detect antibody responses indicating exposure to blood-stage infection within the last 9-months, with 80% sensitivity and 80% specificity [14]. These antigens were down-selected from a starting panel of more than 300 proteins and validation studies of the eight selected proteins were conducted in multiple geographic and epidemiological settings. Isoconazole nitrate Using this data, mathematical Mouse Monoclonal to C-Myc tag modelling demonstrated that with a serological test and treat (PvSeroTAT) regimen, there is the potential to reduce PCR prevalence by 59C69% in endemic areas [14] if anti-hypnozoite treatment with 8-aminoquinolines is provided to those identified as recently infected. This in-development test requires Isoconazole nitrate further optimization and testing of different protein constructs to ensure sensitivity and specificity is maximized, potentially by reducing background cross-reactivity of the antigens in the assay. Previously, two different recombinant proteins of reticulocyte binding protein 2b (PvRBP2b) were identified as the best single antigen predictors of recent infection in a 9-month time frame [14]. Therefore, in the current study the primary objective was to test the performance of multiple fragments within the N-terminal and C-terminal Isoconazole nitrate domains of PvRBP2b.