#	Aladdin ID	Assay Type	Description	Organism	Compounds	Reference	BAO Format	Source
1.	ALA3431446	Functional	GSK_TCAKS: pIC50 Trypanosoma cruzi viability assay using beta-GAL activity. This assay was run at single shot at 5uM compound concentration per well. The biological reagents consisted of NIH-3T3 host cells and trypomastigotes parasitic cells; both cell types were set to a 3.3 * 10^5 cells/mL dilution in assay DMEM by means of a CASY cell counter device using a 60um capillary. 6 uL of the mixture were dispensed per well (about 1 * 10^3 host-cells and 1 * 10^3 parasites) with a Multidrop Combi dispenser. Previously, a solution of 1.67 * 10^5 trypomastigotes per mL in assay DMEM had been prepared and 6uL per well dispensed in the allotted control columns that defined 100% inhibition. The 0% inhibition of parasitic growth control was obtained by leaving the assay mixture untreated in its corresponding plate columns. In all wells, the percentage of DMSO never exceeded 0.5%. Plates incubated for four days at 37C, 5% CO2. The substrate used for the assay fluorescence intensity (FLINT) readout was resorufin-beta-D-galactopiranoside (Sigma-Aldrich) at 5uM per well. It was diluted in PBS supplemented with the soft detergent Igepal (Fluka). Upon addition to the plates, the substrate solution was incubated for 4h at room temperature and the signal read with the EnVision microtiter plate reader (Perkin-Elmer) using the corresponding set of filters (excitation/emission at 531/595nm).	Mus musculus	584	ALA3430912	organism-based format	GSK Kinetoplastid Screening
2.	ALA3431447	Functional	GSK_TCAKS: pIC50 Trypanosoma cruzi Imaging: Infected Cells. H9c2 cells were seeded in T-225 flasks (225cm^2 culture surface; Corning Inc., NY, USA) in DMEM-10% FBS for 4h to allow attachment. Cells were then washed once with PBS before infection. T. cruzi trypomastigotes, collected at days 5 to 8 after infection from LLC-MK2 parasite infected cultures, were allowed to swim out for 4h at 37C from a centrifuged pellet (2,500 rpm/10 min/room temperature). Trypomastigotes were then collected and counted in a CASY Cell Counter. Trypomastigotes, in supplemented DMEM, were added to H9c2 cultures in a multiplicity of infection of 1 and incubated for 18 h. Cells were washed once with PBS before incubation of the infected H9c2 monolayer with trypsin (Life-Technologies) to detach cells from the flask. Cells were counted in a CASY Cell Counter using a 150 um capillary and their density set at 5 * 10^4 cells per mL in supplemented assay DMEM. Infected H9c2 were dispensed into 384-well poly-lysine coated assay plates at 50uL per well using a Multidrop Combi dispenser. After seeding them, the plates were incubated at 37C, 5% CO2 for 72h. Cultures were then fixed and stained by addition of 50 uL of a solution containing 8% formaldehyde and 4 uM DRAQ5 DNA dye (BioStatus, UK) per well. Plates were kept light-protected and imaged one hour later in a Perkin-Elmer Opera microscope using a 20x air objective (NA 0.4) and the following acquisition set: a 635 nm laser excitation line and a 690/50 emission detection filter for DRAQ5 detection. Five images were collected per well for reliable statistical analysis. Automated image analysis was performed with a script developed on Acapella High Content Imaging and Analysis Software (PerkinElmer). Output: percentage of infected cells per well as a second infection marker.	Rattus norvegicus	580	ALA3430912	organism-based format	GSK Kinetoplastid Screening
3.	ALA3431448	Functional	GSK_TCAKS: pIC50 Trypanosoma cruzi Imaging: Amastigotes/Cell. H9c2 cells were seeded in T-225 flasks (225cm^2 culture surface; Corning Inc., NY, USA) in DMEM-10% FBS for 4h to allow attachment. Cells were then washed once with PBS before infection. T. cruzi trypomastigotes, collected at days 5 to 8 after infection from LLC-MK2 parasite infected cultures, were allowed to swim out for 4h at 37C from a centrifuged pellet (2,500 rpm/10 min/room temperature). Trypomastigotes were then collected and counted in a CASY Cell Counter. Trypomastigotes, in supplemented DMEM, were added to H9c2 cultures in a multiplicity of infection of 1 and incubated for 18 h. Cells were washed once with PBS before incubation of the infected H9c2 monolayer with trypsin (Life-Technologies) to detach cells from the flask. Cells were counted in a CASY Cell Counter using a 150 um capillary and their density set at 5 * 10^4 cells per mL in supplemented assay DMEM. Infected H9c2 were dispensed into 384-well poly-lysine coated assay plates at 50uL per well using a Multidrop Combi dispenser. After seeding them, the plates were incubated at 37C, 5% CO2 for 72h. Cultures were then fixed and stained by addition of 50 uL of a solution containing 8% formaldehyde and 4 uM DRAQ5 DNA dye (BioStatus, UK) per well. Plates were kept light-protected and imaged one hour later in a Perkin-Elmer Opera microscope using a 20x air objective (NA 0.4) and the following acquisition set: a 635 nm laser excitation line and a 690/50 emission detection filter for DRAQ5 detection. Five images were collected per well for reliable statistical analysis. Automated image analysis was performed with a script developed on Acapella High Content Imaging and Analysis Software (PerkinElmer). Output: number of amastigotes per cell as infection level measurement.	Rattus norvegicus	580	ALA3430912	organism-based format	GSK Kinetoplastid Screening
4.	ALA3431449	Functional	GSK_TCAKS: pIC50 H2C9 Imaging: H9c2 Host Cardiomyocytes. H9c2 cells were seeded in T-225 flasks (225cm^2 culture surface; Corning Inc., NY, USA) in DMEM-10% FBS for 4h to allow attachment. Cells were then washed once with PBS before infection. T. cruzi trypomastigotes, collected at days 5 to 8 after infection from LLC-MK2 parasite infected cultures, were allowed to swim out for 4h at 37C from a centrifuged pellet (2,500 rpm/10 min/room temperature). Trypomastigotes were then collected and counted in a CASY Cell Counter. Trypomastigotes, in supplemented DMEM, were added to H9c2 cultures in a multiplicity of infection of 1 and incubated for 18 h. Cells were washed once with PBS before incubation of the infected H9c2 monolayer with trypsin (Life-Technologies) to detach cells from the flask. Cells were counted in a CASY Cell Counter using a 150 um capillary and their density set at 5 * 10^4 cells per mL in supplemented assay DMEM. Infected H9c2 were dispensed into 384-well poly-lysine coated assay plates at 50uL per well using a Multidrop Combi dispenser. After seeding them, the plates were incubated at 37C, 5% CO2 for 72h. Cultures were then fixed and stained by addition of 50 uL of a solution containing 8% formaldehyde and 4 uM DRAQ5 DNA dye (BioStatus, UK) per well. Plates were kept light-protected and imaged one hour later in a Perkin-Elmer Opera microscope using a 20x air objective (NA 0.4) and the following acquisition set: a 635 nm laser excitation line and a 690/50 emission detection filter for DRAQ5 detection. Five images were collected per well for reliable statistical analysis. Automated image analysis was performed with a script developed on Acapella High Content Imaging and Analysis Software (PerkinElmer). Output: Number of H9c2 Host Cardiomyocytes nuclei to determine drug-related cytotoxicity.	Rattus norvegicus	580	ALA3430912	organism-based format	GSK Kinetoplastid Screening
5.	ALA3584401	Functional	Antiparasitic activity against Trypanosoma cruzi	Trypanosoma cruzi	1	ALA3580613	organism-based format	Scientific Literature
6.	ALA3583586	Functional	Antitrypanosomal activity against trypomastigote stage of Trypanosoma cruzi Y assessed as inhibition of cell viability after 24 hrs by MTT assay	Trypanosoma cruzi	7	ALA3580607	organism-based format	Scientific Literature
7.	ALA3583589	Functional	Potency index, ratio of benznidazole IC50 to compound IC50 for trypomastigote stage of Trypanosoma cruzi Y	Trypanosoma cruzi	4	ALA3580607	organism-based format	Scientific Literature
8.	ALA3587543	Functional	Antitrypanosomal activity against epimastigote stage of Trypanosoma cruzi CL Brener assessed as growth inhibition after 5 days by MTT assay	Trypanosoma cruzi	15	ALA3585240	assay format	Scientific Literature
9.	ALA3591436	Functional	Antiparasite activity against Trypanosoma cruzi Tulahuen expressing beta-galactosidase assessed as growth inhibition incubated for 96 hrs by CPRG dye based assay	Trypanosoma cruzi	51	ALA3588845	organism-based format	Scientific Literature
10.	ALA3591437	Functional	Antiparasite activity against Trypanosoma cruzi Tulahuen expressing beta-galactosidase assessed as growth inhibition at 10 uM incubated for 96 hrs by CPRG dye based assay	Trypanosoma cruzi	3	ALA3588845	organism-based format	Scientific Literature
11.	ALA3595120	Functional	Antiparasitic activity against Trypanosoma cruzi strain CL Brener epimastigotes after 12 days by ELISA reader	Trypanosoma cruzi	6	ALA3593165	assay format	Scientific Literature
12.	ALA3595121	Functional	Antiparasitic activity against Trypanosoma cruzi strain CL Brener epimastigotes at 300 uM after 12 days by ELISA reader	Trypanosoma cruzi	13	ALA3593165	assay format	Scientific Literature
13.	ALA3595961	Functional	Trypanocidal activity against Trypanosoma cruzi Y bloodstream trypomastigotes in presence of 10% mouse blood assessed as induction of parasite lysis incubated for 24 hrs by Neubauer chamber based cell counting method	Trypanosoma cruzi	9	ALA3593167	organism-based format	Scientific Literature
14.	ALA3598542	Functional	Antitrypanosomal activity against trypomastigotes stage of Trypanosoma cruzi infected in human HT1080 cells assessed as inhibition of parasite growth at 10 uM after 24 hrs by microscopic analysis	Trypanosoma cruzi	7	ALA3596096	organism-based format	Scientific Literature
15.	ALA3598543	Functional	Antitrypanosomal activity against trypomastigotes stage of Trypanosoma cruzi infected in human HT1080 cells assessed as inhibition of parasite growth at 1 uM after 24 hrs by microscopic analysis	Trypanosoma cruzi	3	ALA3596096	organism-based format	Scientific Literature
16.	ALA3598544	Functional	Antitrypanosomal activity against trypomastigotes stage of Trypanosoma cruzi infected in human HT1080 cells assessed as inhibition of parasite growth after 24 hrs by microscopic analysis	Trypanosoma cruzi	21	ALA3596096	organism-based format	Scientific Literature
17.	ALA3602488	Functional	Anti-trypanosomal activity against Trypanosoma cruzi Y epimastigotes assessed as parasite growth inhibition incubated for 72 hrs by MTT assay	Trypanosoma cruzi	10	ALA3600370	organism-based format	Scientific Literature
18.	ALA3606212	Functional	Antitrypanosomal activity against trypomastigote forms of Trypanosoma cruzi Tulahuen C4 expressing LacZ gene infected in rat L6 cells after 96 hrs by inverted microscope analysis	Trypanosoma cruzi	29	ALA3603776	organism-based format	Scientific Literature
19.	ALA3606378	Functional	Anti-parasitic activity against Trypanosoma cruzi Tulahuen C2C4 trypomastigotes infected rat L6 cells incubated for 96 hrs by CPRG/nonidet substrate based inverted microscopy	Trypanosoma cruzi	13	ALA3603826	organism-based format	Scientific Literature
20.	ALA3616328	Functional	Trypanocidal activity against intracellular amastigotes of Trypanosoma cruzi Tulahuen infected in human U937 cells assessed as reduction of viability of intracellular amastigotes by measuring beta-galactosidase activity at 5 ug/ml after 72 hrs by spectrophotometry analysis	Trypanosoma cruzi	2	ALA3612000	organism-based format	Scientific Literature