Detection, quantification and visualisation of protein interactions for hit profiling by high content screening

Alongside established drug target classes, innovative approaches are addressing previously undruggable target classes such as protein–protein interactions. Protein interactions and their relocation within the cell have traditionally been studied using an over-expressed cell model or cell lysate measurements.

This webinar provides an overview of how researchers can use Duolink® which is based on the in situ Proximity Ligation Assay (PLA®) technology, to visualise endogenous protein–protein interactions, in unmodified cells, with unparalleled specificity and sensitivity. For the first time, any target can be readily detected and localized with single molecule resolution by High Content Screening in a 384-well format.

Speakers:

Dr. Bernhard Ellinger (Principal Scientist, European ScreeningPort)
Proximity ligation assays for Hit Profiling in small molecule drug discovery campaigns – a case study

Thomas Juehne (Principal Scientist, Sigma-Aldrich)
Visualising endogenous protein interactions and translocations


Topics covered:

  • Compound Profiling using PLA® as part of the hit-to-lead workflow.
  • Direct in cell visualization of endogenously expressed proteins and their interacting partners.
  • The ability to provide evidence for target engagement of the compound.

Dr. Bernhard Ellinger (Principal Scientist, European ScreeningPort)
Proximity ligation assays for Hit Profiling in small molecule drug discovery campaigns – a case study

Proximity ligation is a highly specific and sensitive technology to monitor individual proteins, their interactions and post-translational modifications in situ. It provides single molecule resolution and objective and unbiased quantification in cells and tissues on endogenous expression levels. The European ScreeningPort has optimized the PLA® technology for use in an automated 384-well screening format and has also implemented routines for automated image analysis including spot detection, localisation and quantification. In this presentation, we will present a case study of a small molecule drug discovery project performed in collaboration with a biotech partner. The target mechanism involved the disruption of the interaction between amyloid ß oligomers and RAGE (Receptor for Advanced Glycation Endproducts). Profiling of compounds during Hit-to-Lead stages has involved a variety of High Content functional cellular assays including PLA. We will discuss the use of PLA in 384-well format as part of a Hit-to-Lead screening cascade to monitor compound-mediated disruption of this receptor-ligand complex.
 
With a background in Biochemistry Bernhard Ellinger did his Ph D in Chemical Biology in the lab of Herbert Waldmann (Max Planck Institute of Molecular Physiology, Dortmund). At European ScreeningPort GmbH he is primarily involved in high throughput and high content screening and image data analysis. His core expertise lies in biochemical as well as cellular assays with a focus on label-free formats and imaging techniques. He has >3 years of experience in assay development for proximity ligation technology.

Thomas Juehne (Principal Scientist, Sigma-Aldrich)
Visualising endogenous protein interactions and translocations

Insight into protein pathways is critical to the drug discovery process. Protein interactions and their relocation within the cell have traditionally been studied using an over-expressed cell model or cell lysate measurements. We will present Duolink®, which is based on the in situ PLA® technology, a method that allows the direct in cell visualization of endogenously expressed proteins and their interacting partners
 
Tom has over 30 years of diverse and extensive research experience. Trained as an organic chemist, he has been active in life science research for the past 25 years. While at Sigma-Aldrich Tom has been engaged in several technologies. Examples include: surface modification chemistry, conjugation methodologies, bioactive nanomaterials, labeling, detection, imaging and site-specific conjugation methodologies.