Services Offered



I. Sample Preparation


Affinity Purification (FLAG Epitope)

This assay will identify interacting protein partners (prey) for a protein of interest (bait). These assays can be performed reiteratively with baits from the same pathway or protein family to create intricate and detailed maps of signalling networks. We will provide the appropriate vector containing a Triple-FLAG epitope for the generation of stable cell lines. Once stable cell lines are generated expressing the baits of interest, we will perform affinity purification from cell pellets by lysing the cells, purifying the bait(s) and interacting proteins using anti-FLAG magnetic beads, and digesting the protein into peptides in preparation for mass spectrometry analysis.


Affinity Purification (BioID)

This assay will identify proteins in close proximity to a protein of interest (bait) within the cell. The bait is tagged with a promiscuous birA that adds a biotin moiety to proteins in close proximity. The biotinylated proteins are then captured with streptavidin beads and identified by mass spectrometry. We will provide the appropriate birA epitope-tagged vector for the generation of cell lines. Once stable cell lines are generated expressing the baits of interest, we will perform affinity purification from cell pellets by lysing the cells, purifying the biotinylated proteins using streptavidin-coated magnetic beads, and digesting the protein into peptides in preparation for mass spectrometry analysis.


Customized Affinity Purification for Mass Spectrometry

In addition to our two standardized services, we also offer custom affinity purifications. These services include affinity purification of an endogenous protein by its cognate antibody or purification of proteins by immobilized DNA, peptide, protein or small molecule. In each case, the cost will be determined on a per sample basis calculated after discussion of project design and will include assay setup charges.



II. Mass Spectrometry Analysis

We offer a range of mass spectrometry analyses on high-resolution, high-sensitivity mass spectrometers. The first step in mass spectrometry-based proteomics analysis is the identification of proteins within the sample through discovery analysis using an approach termed 'data-dependent acquisition'. If desired, we also offer additional services to quantify the relevant amount of proteins within samples using either comprehensive or targeted approaches.


Discovery Analysis (Data-Dependent Acquisition (DDA))

Digested protein samples will be analyzed with a high-resolution, high-sensitivity mass spectrometer in a nano-LCMS/MS workflow. Data will be stored in a secure manner within the ProHits mass spectrometry LIMS system, and analyzed with leading search engines and algorithms. In the case of samples for identification, a confident list of protein identifications with peptide details will be reported. In the case of protein-protein interaction studies, the report will include a list of significant interacting proteins (with assigned pValues from the SAINT algorithm) in formats compatible for visualization (e.g. through cytoscape).


Comprehensive Quantitative Analysis - Data-Independent Acquisition (DIA-SWATH)

For projects where discovery proteomics has been successful and there is a desire to quantify changes in protein composition under different conditions, we will perform relative quantitative analysis. Unlabeled digested protein samples will be analyzed using a comprehensive quantification method (DIA-SWATH) that determines the relative intensity for all peptides above a signal-to-noise threshold within the mixture.


Targeted Quantitative Analysis

For quantification of specific set of proteins, peptides or post-translational modifications, we offer targeted quantification analysis (MRM-HR or Targeted SWATH on a Triple TOF mass spectrometer). The costing will be determined after the project is defined and will include all assay setup costs.


Labelled Quantitative Analysis

We also offer the ability to multiplex samples for quantification using SILAC, iTRAQ, and di-methyl labelling. Please contact us to discuss your project and the most appropriate methodology.


Data Analysis

For customers who wish more detailed data analysis or our assistance in integrating proteomics data with other data, we offer extended data analysis from our mass spectrometry specialists or our in house statistician.