High Throughput Screening Laboratory
The HTS laboratory has all of the equipment required for HTS. The laboratory operates two Beckman FX liquid handlers, a Beckman SPAN-8 liquid handler for cherry picking, eight Multidrop instruments for dispensing reagents into plates, two plate washers, two plate sealers, two EnVision multimode plate reader, a Coulter Cell Counter and all necessary cell culture equipment. The laboratory has a DB Pathways 855 with cell incubator and robotics for microscope based screening.
The HTS Laboratory has conducted two compound screens and two genomic siRNA screens simultaneously, representing the current maximum daily screening capacity of the laboratory. Individual experiments differ considerably in throughput, depending upon assay complexity. Typical screening rates are 25 to 40 plates per day for cell based compound screens (8,000 to 12,800 compounds) and 10 siRNA source plates containing 800 gene specific sequences for RNAi.
The latter experiments usually require comparing two conditions in triplicate (60 plates per day). At these rates HTS of either compounds or siRNAs requires five weeks for primary screening and one to two weeks for repeating failed plates, cherry picking, and hit validation.
In 2006 the HTS lab completed five compound screens and seven genomic siRNA screens, with two additional compound screens in progress at year’s end. Typically, two to four assays are being developed for HTS at any time, with some of these not proceeding to full HTS for lack of funding.
The laboratory currently could conduct as many as 20 compound or siRNA experiments per year. Lack of funds for HTS experiments rather than limitations of infrastructure is the major factor restraining the growth of such experiments at UT Southwestern.
A major activity of the HTS lab during assay development is to minimize screening costs by limiting the cost of reagents. The most common compound or siRNA assay run by the laboratory is cell-based with a luciferase reporter. Reagent costs for HTS experiments conducted in 2007-2008 ranged between $0.25 and $0.06/well with the lower figure the target for assay development.
A screen of the full compound library usually requires 700 plates, including those for repeating failed plates and assay development. Plate and liquid handler tips are approximately $7,000-$10,000/screen and regent costs at $0.15 per well are approximately $40,000. Screens of the human genomic library typically require $20,000 in reagent costs.
In addition to assay reagents, there are modest costs for preparing compound or siRNA assay plates that are charged to the laboratory initiating a screen and certain equipment has user fees to support service contracts (see list of fees below). The major costs for the experiments, the expensive equipment, the cost of acquiring and formatting the compound and siRNA libraries and staff salaries are shared by the NCI program project, the Cancer Center, and UT Southwestern.
Tissue culture can be conducted in the investigator’s laboratory or the HTS laboratory. If long term culture work is to be conducted in the HTS lab, a daily fee of $5 will be assessed for use of HTS pipets, tubes, gloves, paper towels, and alcohol. The investigator’s laboratory should supply their own plates or dishes, media, bottles, and any filter units needed.
For all RNAi screens, a supply fee of $425 will be assessed. This is to cover the cost of tips ($250.00), plate seals ($120.00), and alcohol ($55). siRNA oligos are supplied free to investigators whose projects have been approved. For all microRNA screens, a supply fee of $100 will be assessed. This is to cover the cost of tips, ethanol, tubes, pipettes, plate sealing film, and the robot service contract.
Investigators using the high content BD pathway screening microscope must pay $5/hour for use of the microscope. Data analysis can be conducted on the Attovision Workstation free of charge. This charge is to cover the cost of the microscope service contract.