**1.1** *T. cruzi* **infection diagnostic during the different stages of the illness**

The infection diagnosis is not a simple task because the features of the illness development as well as the immunological response of the host must be taken into account when performing the laboratory diagnosis. Different diagnostic methods are used, depending on the illness stage and the particular clinical entity of the patient.

The alternative ways to get acute Chagas' infection are mainly vectorial and vertical transmission, and the reactivation of the chronic disease in immunosuppressed individuals. Therefore, diagnosis at this stage limits to: *i)* the uncommon cases of symptomatic patients, *ii)* newborns delivered from chagasic mothers, and *iii)* immunosuppressed patients that have been previously diagnosed as infected.

In the above mentioned cases, parasites are usually, easily found in blood, therefore being their direct microscopic observation the chosen method for a safe diagnosis.(WHO, 2002;Rosenblatt, 2009) In contrast, indirect serological techniques present low sensitivity in recently infected individuals because humoral immune response is delayed.(Zuniga *et al.*, 2000) Moreover, even specific IgM immunoglobulin has a window period that requires up to several weeks to be produced at detectable concentrations. Other pitfalls such as the lack of IgM anti-*T. cruzi* commercial kits prevents from choosing serology as the infection diagnostic method. This methodology is only used when the parasite is not found in fresh blood smears, by Struot and/or microhematocrit methods.(Luquetti & Schmunis, 2010)

During the chronic phase, the *T. cruzi* infection diagnosis is required, *i)* as pregnancy or occupational routine control analysis -in countries where according legislation is in force-, *ii)*  when in presence of a Chagas' compatible cardiopathy, *iii)* during transfusion and transplantation screening, and *iv)* to achieve a reliable illness prognosis of patients who were already diagnosed. One particular case among the latter ones is monitoring the antiparasitic treatment effectiveness.

Conversely to what happens in the acute phase, during the chronic infection, a significant humoral response is found in immunocompetent individuals, along with low parasite concentration in blood. Under these circumstances, indirect techniques are highly sensitive, while the direct ones fail. Hence, Chagas' disease serologic diagnoses intend to verify the occurrence of several specific antibodies against *T. cruzi*. However, conventional serology presents different sensitivity and selectivity, depending on the immunological technique used to perform the determination, and mainly in the nature of the antigen used to capture the specific antibodies.(Belluzo *et al.*, 2011)

In this Chapter, we will describe the most recent contributions of our and other groups to improve the analytical tools available to properly and safely diagnose Chagas' disease using recombinant proteins, in each one of the clinical entities mentioned above.
