Screening Services

The Yale Center for High Throughput Cell Biology offers comprehensive screening services for clients wishing to run full or partial genome screens. Assay can be designed for imaged based high content analysis using the PerkinElmer Opera or for simple fluorescence or luminescence readouts using our Tecan InfiniTe M1000 plate readers. Beginning with the initial assay development, our staff will work with you through the primary screening campaign, confirmation of hits and final informatics analysis of the results. Our services are designed to be as modular as possible, allowing investigators to choose from several options. More information on the level of services can be found below or by visiting the individual sites. In all cases, the client is responsible for the costs associated with assay specific supplies (i.e., antibodies, media, supplements and any additional reagents).

Assay Development
The assay development portion of the screen is typically run as a collaboration between the HTCB and the referring investigator. All experiments can be run on a fee for service basis where investigators can choose their level of interaction with the HTCB. Plates can be run independently and supplied to us for imaging or members of your lab can choose to run the experiments at our facility. We offer lab space, cell culture facilities and a full suite of equipment designed to facilitate work in microwell plates. In addition, investigators can choose to transfer the development work to our scientists, with results provided at the end of each experiment. Clients choosing the modular approach will be billed on an hourly rate for the equipment used. Those who choose to have the work done by our staff will be charged a fee for each experiment.
We have designed a series of steps to help ensure each screening campaign will be successful.
• Choosing and testing an assay readout
• Optimizing the biological system (cell line and culture conditions)
• Determination of dynamic range and variation in the assay
• Transfection Efficiency Testing (for cell lines not previously used by the HTCB)
• Determination of suitable controls (both positive and negative)
• Generation of plate level statistics for the assay (Z and Z’ Factors)

Screening
Upon the conclusion of assay development, the assay will be transferred to our staff for all further steps through the completion of the primary screen. Before beginning the screen, a series of final optimization experiments will be performed by us, culminating in a small-scale validation screen. The details of these steps are listed below.

Assay Optimization
Assay Optimization is designed to allow our scientists to test the final conditions of the assay. Making sure the cells and other reagents perform in a manner consistent with the development phase of the project is essential prior to screening. This step serves to:
• Familiarize our staff with handling the assay
• Ensure functionality and proper settings of all equipment
• Test for optimal transfection efficiency
• Test for even cell density across a plate
• Ensure low variation between wells

Validation
The validation screen consists of 3 plates representing 718 genes from the kinase portion of the library, run in triplicate (10 plates total, including a control). These plates are created and arrayed in the same way as the genome wide library to replicate the exact conditions of the full screen. At the end of the validation a meeting will be scheduled to discuss the performance of the assay before continuing and provide a data report for the experiment. The data report will address:
• Performance of the controls across multiple plates
• Plate level statistics (Z and Z’ factors, if applicable)
• Data normalization (Percent inhibition, z score, robust z score)
• Analysis algorithm optimization (imaging assays only)
• Correlation between replicates
• Screening artifacts (i.e., edge effects)
• Activity including a preliminary hit rate


A representation of a typical screening plate can be found on the left. Each plate contains a maximum of 320 samples arranged in the middle of the plate. Positive and negative controls are arranged in a checkerboard pattern in columns 2 and 23, with 16 wells of each control per plate. Columns 1 and 24 are reserved for any additional control or conditions that may be needed for the assay.


Primary Screen We offer full and partial genome wide screens using the Dharmacon human siGenome collection. The full library consists of over 18,000 genes arrayed across 58 plates. We recommend that each screen be run in triplicate to limit the potential for false positives and negatives, however investigators wishing to run fewer replicates can do so at their own discretion.
Clients wishing to run subsets of the library can choose from one or more of the pre-arranged sets that can be found on our library page. Our scientists will be responsible for carrying out the genome wide screen, except in cases where a specialized step may require assistance from a member of your lab.
Typically, a screen is broken down into smaller batches run over several weeks. The size of each batch and the time until completion depends upon the assay. Included in each batch is a control plate to test for variability. A full screen in triplicate consists of roughly 180 plates, including controls.