full issue - Association of Biotechnology and Pharmacy

Current Trends in Biotechnology and PharmacyVol. 5 (2) 1130-1133 April 2011. ISSN 0973-8916 (Print), 2230-7303 (Online)Automated Synchronization of P. falciparum using aTemperature Cycling Incubator1130Alejandro Almanza, 1, 2 Lorena Coronado, 1 Nicole Tayler, 1 Liuris Herrera 1, 2 and1, 2*Carmenza Spadafora1Centro de Biología Celular y Molecular de las Enfermedades (CBCME), Instituto de Investigaciones Científicas yServicios de Alta Tecnología (INDICASAT AIP), Panama.2International Cooperative Biodiversity Groups (ICBG), Smithsonian Tropical Research Institute (STRI), Panama.*For Correspondence - cspadafora@indicasat.org.paAbstractAs malaria keeps affecting millions of livesevery year, research based in culture ofPlasmodium falciparum in vitro needs to beefficient and accurate. The development of bettertechniques and methodologies for the growth andmaintenance of the parasites can save money,time, and lead to more trustable results. It hasbeen observed, first in patients and then in thelaboratory, that the malaria falciparum parasitesgrowth is affected by high temperatures. This traitcan be used with laboratory cultures tosynchronize and maintain the parasites in the samestage of their cell cycle. This harmony of stagesis very desirable for the purpose of conductingmetabolomic, proteomic and transcriptomeanalysis as well as for drug screening. Mostscientists in the field of malaria use chemicals(usually sorbitol) that kill certain stages of theparasite to obtain synchronization, but this lattermethod does not last long and the parasites thustreated should not be used for assays immediatelyafter the treatment, due to the toxic effects thatmight have been infringed in the culture. Atemperature cycling incubator (TCI) was acquiredin our laboratory and it was used to test thesynchronization of the multidrug resistant W2 andchloroquine resistant 7G8 strains, commonly usedin our bioassays where they and theirsynchronization constitute essential tools for ourdrug discovery program. We followed the protocoldesigned by Haynes and Moch in 2002 and wemade a comparison of the effectiveness of eachof the two methods, chemical and temperaturebased. Our results show W2 synchronization bytemperature cycling, with the help of an initialuse of 0.3 M alanine, to last more than two monthswhile a tight synchronization with the use of 5%sorbitol was lost as rapidly as in one week. Sorbitolcould also be used with the TCI forsynchronization with good results. However, 7G8could not be efficiently synchronized withtemperature cycling using the same program asthat of W2.Key Words: P. falciparum, Culture,Synchronization, High temperature, TCI.IntroductionIt has been described that in vitro growthof Plasmodium falciparum is inhibited by hightemperatures (1-5) and there have beenrelationships drawn from these findings on howfever in malaria patients may influence parasitegrowth, coordination of cyclic stages, andparasitemia.In 2002, Haynes and Moch published amethod for using these observations tosynchronize P. falciparum cultures for laboratoryAutomated Synchronization of P. falciparum