In this work, we propose a methodology to study microorganisms chemotaxis in real time using an Optical Tweezers system. Optical Tweezers allowed real time measurements of the force vectors, strength and direction, of living parasites under chemical or other kinds of gradients. This seems to be the ideal tool to perform observations of taxis response of cells and microorganisms with high sensitivity to capture instantaneous responses to a given stimulus. Forces involved in the movement of unicellular parasites are very small, in the femto-pico-Newton range, about the same order of magnitude of the forces generated in an Optical Tweezers. We applied this methodology to investigate the Leishmania amazonensis (L. amazonensis) and Trypanossoma cruzi (T. cruzi) under distinct situations. ©2009 Optical Society of America.7367 http://www.who.int/tdr/diseases/chagas/diseaseinfo.htmlWorld Health Organization. 2002. Annex 3: Burden of disease in DALYs by cause, sex and mortality stratum in WHO regions, estimates for 2001. The World Health report. Geneva: WHO 2002: 192-197Fontes, A., Giorgio, S., de Castro Jr., A.B., Neto, V.M., Pozzo, L.Y., Marques, G.P., Barbosa, L.C., Cesar, C.L., Determination of femtonewton forces and fluid viscosity using optical tweezers: Application to Leishmania amazonensis (2005) Proceedings of SPIE, 5699, pp. 419-425Alves, C.R., Albuquerque-Cunha, J.M., Mello, C.B., Garcia d, E.S., Nogueira, N.F., Bourguingnon, S.C., de Souza, W., Gonzalez, M.S., Trypanosoma cruzi: Attachment to perimicrovillar membrane glycoproteins of Rhodnius prolixus (2007) Experimental Parasitology, 116, pp. 44-52Cortez, M.G.R., Gonzalez, M.S., Cabral, M.M.O., Garcia, E.S., Azambuja, P., Dynamic development of Trypanosoma cruzi in Rhodnius prolixus: Role of decapitation and ecdysone therapy (2002) ParasitologyResearch, 88, pp. 697-703Kohidai, L., Chemotaxis: The proper physiological response to evaluate phylogeny of signal molecules (1999) Acta Biol Hung, 50, pp. 375-394Bagorda, A., Parent, C.A., Eukaryotic chemotaxis at a glance (2008) J. Cell Science, 121, pp. 2621-2624www.biophysics.org/education/williams.pdf