Small Compound Screening Overview

The initial consultation with the HTS facility manager is an opportunity to discuss your ideas for a potential screen during the conceptual stage or as you begin the bench-top development.

We have experience across a diverse range of screening platforms and formats and have run a wide variety of screens in our facility. We are able to advise you on what might be the most effective approach (both in terms of time and cost) for the question you would like to investigate.

We have also developed a number of novel screening approaches designed to answer specific criteria and we welcome the opportunity to develop additional novel screens. We would encourage you to discuss your proposed assay at the earliest stages of development as we can assist you with assay set-up, choice of reagents, specific phenotype optimization, logistics, timing and projected costs. 

HTS Transfer protocols

All assays being transferred to the Janus for HTS will be subjected to a thorough validation process before the production screening beings.

This validation process will consist of:

• Initial Consultation
• Stability and Process study
• Liquid Handling Validation
• Plate Uniformity Assessment
• Control Validation, Z'calculation, Assessment of edge effects and drift within the body of the screen
• Replicate Experiment - We require as a minimum a 2 replicate study over 2 different days for biological reproducibility and robustness
• Pilot Screen - a small number of plates containing compounds of varied pharmacologically activity and the chosen controls
• Production Runs

CRISPR/Cas 9 Cell Screening Facility

CRISPR/Cas9 technology is ideally suited for genome-wide screening applications due to the ease of generating guide RNAs (gRNAs) and the versatility of Cas9 or Cas9 derivatives to knockout, repress, or activate expression of target genes. Several pooled lentiviral CRISPR libraries have been developed and are now publicly available. Here at the TDI we have both Pooled Lentiviral CRISPR genome knockout and CRISPR gain-of-function libraries.

Knock-down and activation libraries

The CRISPR/Cas9 system is ideal for genome-wide knockout and gain of function screening experiments due to the ease of generating gRNAs and the efficiency and irreversibility of Cas9-mediated genetic modifications. Here at the TDI we have pooled lentivirus CRISPR knock-out and activation libraries that target all the genes in human and mouse genome.

Table 1.  List of CRISPR library 

Virus production

We have the facility to produce lentivirus for pooled CRISPR lentivirus and provide advice (Figure 2). Most of the CRSPR libraries in table 1 have been used to make Lentivirus and is available for use in screenings. 

 Figure 2. Schematic diagram of Lentivirus production

Screening and sequencing

The aim of the facility is to provide assistance in all aspects of lentiviral CRSPR screening from initial experimental design, virus production, purification, transduction, screening and  FACS sorting.   We can perform FACS sorting of the pooled genome-wide CRISPR screens using the benchtop SH800 cell sorter. SH800 has a 488nm excitation laser and permits sorting of a wide range of cell sizes and applications using different microfluidics sorting chips (70 μm, 100 μm, and 130 μm). 

For sequencing, we can help and assist in genomic DNA extraction and genomic PCR to prepare samples for sequencing. We have PCR primers for all of the CRISPR libraries and the technical resources and knowledge.