Cutting-edge proteomics technologies
Single cell proteomics
While single cell techniques in genomics are widely used, methods for single cell analysis in proteomics are still being developed. How the expressed proteome differs from cell to cell is a critical question, which complements the question addressed by genomics.
Mass spectrometry - the state-of-the-art technique in proteomics - can detect and quantify nearly complete and quantified proteomes, starting from tens of thousands of cells or more. Conversely, the technique of mass cytometry makes it possible to quantify proteins, but only in small numbers and on proteins that are sufficiently abundant.
In a typical mammalian cells, reliable analysis by mass spectrometry was only possible for the most abundant proteins. New approaches may soon bring about changes.
Nanopore sensors, a now well-established technology for sequencing DNA and RNA at the level of individual cells, are also being investigated for protein sequencing.
Immuno (antibody detection) approaches for protein profiling in single cells are also increasingly being applied on a larger scale.
Targeted proteomics is a general method of detecting a set of proteins of interest. However, the few years of practice have given way to great disparities in approaches, notably by combining several instruments (quadrupoles, orbitrop, MALDI-ToF). Several studies have yielded very good results.
The four methods as classified by Eva Borràs et Eduard Sabidò are the following:
Selected ion monitoring (SIM): this involves filtering the spectrometer detector for a single peptide, which is used for quantification. The quantification is done in detection per second and can be done sequentially on several peptides.
Targeted analysis at the peptide level (MS1): The data acquisition is done for all the peptides present in solution over the total duration of the color gradient, which creates an MS1-Map (MS1-Map).
Targeted analysis at ion fragment level (MS2): One of the major strategies of targeted proteomics, the Selected reaction monitoring (SRM) uses the triple quadrupole MS with a filter on the peptides to be monitored, which will then be analyzed by another MS analyzer. This strategy has a relatively low background noise and high sensitivity.
Parallel reaction monitoring (PRM): simultaneous analysis of several peptide fragments pre-selected by a quadrupole to isolate the peptide of interest, then a MS in screening mode.
Discovery proteomics is the study and discovery of markers for a given test by mass spectrometry.
The number of samples initially tested is relatively small.
Once these markers are identified, they are used on a larger scale on a larger number of samples. Advances in discovery are mainly related to the improvement of analytical instruments and software.
Types of providers
Two major categories of proteomics providers:
Academic platforms are solicited for proteomics requiring the use of expensive equipment or requiring method development. Mass spectrometers are very difficult to finance, which academic structures can do by grouping together. The service is provided by a technological platform available from external structures such as research institutes and companies.
The service companies
Proteomics service companies offer services adapted to the demands of private or public clients: identification, quantification, method development for clinical tests. They can be highly specialized or generalist and can validate methods using GLP (good laboratory practice) principles.
Clinical and diagnostic applications of proteomics
The application of protein (or peptide) biomarkers in clinical studies is a dynamic and rapidly growing field. The introduction of clinical proteomics, such as mass spectrometry-based assays and antibody-based multiplexed protein arrays, has reshaped the landscape of biomarker identification and validation, enabling the discovery of new biomarkers at an unprecedented rate and with unprecedented reliability.
The methods used are typically quantitative mass spectrometry (label-free or SILAC / iTRAQ), capillary electrophoresis coupled to MS (ESI-MS),
The research fields in which proteomic analyses are in progress are the following ones:
Proteomic Analysis in Oncology
Many markers are used in oncology, as is the case for an sponsored by the Montpellier Cancer Institute, which measures five proteins present in the blood (adenylate kinase 2 (AKongoing clinical study2), isocitrate dehydrogenase 2 (IDH2), annexin 1 (ANX1), DNA-apurinic or apyrimidinic site lyase (APEX1), and heat shock cognate 71 kDa (HSC70) ), and which will allow to stratify patient groups and recommend treatments accordingly.
This example is representative of the use of existing treatments at different doses for different patient subgroups, and holds promise for personalized medicine, which can limit certain side effects and be more cost-effective.
Proteomic analysis in nephrological dysfunction
Markers present in urine can be identified and measured by proteomic techniques, as is the case for patients with type 2 diabetes and at risk of developing kidney disease. In a study conducted by the Steno Diabetes Center Copenhagen which has been finalized, a cocktail of peptides (derived from inflammatory proteins, fibrinogens, apolipoproteins, alpha-1 antitrypsin) is analyzed by EC-MS in the urine of 1777 patients, and a standard treatment vs sprironolactone vs placebo is administered randomly.
Proteomic analysis for inflammatory diseases
A classic protein marker of inflammation, C-reactive protein (CRP), is measured with the aim of stratifying subgroups in a cohort of patients under intensive medical observation for persistent infections.
Proteomic analysis for neurological diseases
The S100B protein is a standard biomarker of brain damage and has the advantage of being detected non-invasively since it is found in the blood after blood-brain barrier dysfunction.
This marker is at the heart of several studies, notably in parallel with the study of other markers such as P and Y neuropeptides, neurofilament triplet protein (NFL), or in the context of head trauma follow-up in comparison with head CT scans.
Proteomic analysis for cardiovascular disease
In the case of cardiovascular diseases, a precise and rapid diagnosis is essential. Several ongoing studies are using proteomics to identify different markers, such as the Fatty Acid Binding Protein (FABP), or like this other study that looks at 16 different cardio markers at once (apolipoprotein B100, FABP3, L-selectin, atrionatri-uretic peptide receptor 1, insulin-like growth factor-binding protein 3, coagulation factor V [F5], F9, F10, adiponutrin, von Wille-brand factor, thrombospondin 1, prolactin, paraoxonase 3, epidermal growth factor receptor, vascular endothelial growth factor, henopexin, myeloblastin, plasma serine protease inhibitor, heparin cofactor II, hyaluronan-binding protein).