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High-Throughput Bioscience Center

Who We Are

HTB Team
The High Throughput Bioscience Center (HTB) consists of members with advanced training in biological science, chemical science, and engineering. We have professional experience in target identification and validation, assay development, high throughput bioscience, laboratory automation, and management of scientific collaborations.

What We Do

  1. Assist St. Jude principal investigators (PIs) in assay development
  2. Validate and automate assays
  3. Implement and manage HTB projects
  4. Coordinate and implement secondary assays; provide SAR biology supports
  5. Utilize and assist the management of compounds in conjunction with other Centers within the Chemical Biology and Therapeutics department.
  6. Manage and utilize whole genome siRNA libraries (human and mouse)
  7. Evaluate and develop novel drug discovery technologies
  8. Provide support letters and information to assist in grant applications for St. Jude PIs
  9. Provide training to the St. Jude community (e.g., technology, process)

How We Collaborate

The HTB process can be represented by this overview:HTB Process

  • Determination of assay type. We are capable of performing biochemical/enzymatic assays, as well as cell-based assays. Commonly, assays use read-outs such as absorbance, luminescence, or fluorescence measured by a multi-well plate reader. High content (image-based) assays are also available.
  • Miniaturization of the assay. 384-well plates are the current standard for screening at St. Jude. To aid the optimization process, we are happy to provide collaborators with a few of the plates we routinely use during screening.
  • Assay simplification. The goal is to achieve a successful assay in the fewest number of possible steps. Often this requires combining multiple steps (for instance by pooling reagents) or removing steps that are not essential.
  • Assay reproducibility. To compare data points within the enormous screening datasets, it is imperative that the assay performs reliably throughout the screening campaign. Reproducibility is often linked to reagent quality and stability, so reagent stability must be verified for the expected run time of the screen (typically 6-10 hours). Each screening run is subjected to data quality inspection using widely accepted quality metrics such as Z'-factor or SSMD.
  • Assay signal window. The goal is to have good separation between positive and negative controls. This requires a large readout window between positive and negative controls, as well as low variance within each control. For a normally distributed data set with appropriate positive and negative controls, Z'-factor is a good, simple measure of assay quality.

Assay development is the responsibility of labs and individuals interested in collaborating with us. Our staff is happy to offer advice and assistance when possible. We highly recommend that new collaborators communicate with us early on in the assay development process so we can provide appropriate guidance. 

Assay Validation

During assay validation, the quality and feasibility of the assay are verified for the purposes of automation. The two primary goals during assay validation are:

  • Validation of the assay itself. An assigned HTB member will reproduce the assay to determine the assay robustness. A number of statistical tests will be applied to the data to ensure that the assay quality is high enough to justify the cost of time, people, and other resources required to formally initiate a screen.
  • Validation of the automation. The assay will be tested on the robotic screening decks to ensure that the robots can reliably perform the assay. Since precise timing is critical for many assay types, timing analysis will be performed at this stage to establish that all plates are handled identically.

Validated assays have a high probability of success in a high throughput screen, but until screening begins it is impossible to predict every possible complication. Without assay validation, there is no reason to run the screen, since the results would not be trustworthy.

The final stage of assay validation is the completion of an assay validation report. This document serves as an important record of the assay design, reagents used, assay protocol, automation protocol, and statistical analysis of the data acquired during the assay validation process. Collaborators work with the HTB Center and other Chemical Biology and Therapeutics (CBT) Centers to ensure that this document is accurately completed prior to the primary screen.

Primary screen

  • Small-molecule screens: The validated assay is tested against CBT's small molecule library at a single concentration. 
  • siRNA screens: Primary siRNA screens are performed with pooled siRNAs, where each well contains four siRNAs.

Most assays operate with a throughput of 60-100 plates per day. Depending on the assay, the collaborator may be highly involved during the primary screen (for instance to prepare reagents or cells for the run), particularly at the beginning and end of screening runs. The data from each screening run are subjected to a quality control check using in-house data analysis software (e.g., RISE, GUItars). Problems in data quality/consistency will require screening to halt until the problems are resolved.

Hit selection

The primary screen is complete when the desired number of compounds/siRNAs are screened, assay quality is maintained, and "hits" are selected. Hit selection criteria are determined after discussion among the project team members (e.g., the collaborator, members of the HTB Center, a CBT chemist, and Lead Discovery Informatics).

Secondary screening

  • Small-molecule screens: Following the primary screen, "hit" compounds are cherry-picked from the compound storage facility and arrayed in serial dilutions on new compound screening plates. These dose-response (DR) plates are generally used for all secondary screens. First, the same assay used in the primary screen is repeated in triplicate with the DR plates to validate the activity of the compounds and to determine the potency of the compounds. Next, the DR plates are used in other secondary assays. Useful secondary assays can establish hit specificity, reveal undesired cytotoxic effects, and/or confirm activity through an orthogonal assay of the desired bioactivity.
  • siRNA screens: The primary screen is followed by a confirmatory screen, where "hits" are tested in quadruplicate. Once the "hits" are confirmed, a deconvolution screen is conducted in quadruplicate using individual siRNA in each well.

Resource Management

Full-Time Equivalent (FTE)

  • Assay development typically occurs in the individual PI's lab with input from the HTB Center.
  • Assay validation, automation, and screening implementation will be performed at the HTB Center with designated FTEs from both the HTB Center and the individual PI's lab.
  • Lead evaluation and optimization typically occurs at CBT and involves multiple Centers.
  • CBT informatics and chemistry provide support for screening and post-screening efforts, where applicable

Cost

  • Currently, CBT is responsible for compound supply, infrastructure maintenance, and general reagents; a charge-back system may be established according to institutional guidelines in the future.
  • Individual investigators are responsible for study-specific reagents.