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Protein Identification Services

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The proteins are direct products of genome, they are intricate biomolecules and macromolecules in organism. The structure of proteins is composed of one or more elongated polypeptide chains, while peptides are formed by amino acid residues. And each amino acid possessing a distinct mass and each protein exhibiting its unique functionality. The liquid chromatography tandem mass spectrometry (LC-MS/MS) technique is characterized by its exceptional sensitivity, accuracy, and automation capabilities, enabling precise determination of peptides and proteins with relative molecular masses. Additionally, it allows for the identification of amino acid compositions and detection of post-translational modifications (PTMs). The LC-MS/MS technology is extensively employed for the quantitative and qualitative analysis of proteins. Importantly, the application of MS-based protein identification is crucial in the investigation of major diseases and pharmacological control mechanisms.

Protein identification is the primary focus of proteomics related analyses. The current protein identification strategies primarily include bottom-up, middle-up, and top-down approaches, and with the bottom-up method being the most widely employed. A typical bottom-up proteomics workflow consists of several major steps: (i) extraction of protein mixture or purification single protein from biological samples, followed by (ii) protein concentration determination (e.g., bicinchoninic acid assay). Then (iii) the proteins are proteolytically cleaved by proteases (trypsin or lys-C typically) and (iv) analyzed by LC-MS/MS. (v) The resulted MS data files will undergo protein identification through database search software and be subjected to analysis using bioinformatics technology. Furthermore, the utilization of protein fractionation techniques such as SDS-PAGE and chromatographic columns, or peptide fractionation techniques like reversed-phase liquid chromatography, facilitates enhanced high-throughput analysis and increased depth in coverage. The field also encompasses a wide range of peptide enrichment techniques for the identification of diverse PTMs in proteins.

Protein identification serviceFigure 1. Illustration of three approaches for protein identification and characterization.

Our protein identification services

Creative Proteomics accept all types of samples (e.g., gel spots, gel bands, solution samples, cell, tissue, body fluid) and will provide high sensitivity protein identification service by using the latest LC-MS technologies. For protein identification analysis, 1) We can provide assistance in determining the purity, quantity, and identity of proteins. 2) Additionally, we offer expertise in analyzing protein expression and localization. 3) Our services also encompass the investigation of PTMs in proteins. 4) Furthermore, we specialize in studying protein induction and turnover. 5) We are also expert in investigating protein's interacting partners and networks through IP/Pull-Down protein samples.

We have developed advanced technologies and accumulated over ten years' extensive expertise in protein identification. Equipped with experienced technical team, stringent quality control system, and advanced LC-MS platform, we could deliver the reliable and accurate protein identification data to you.

Creative Proteomics can provide a variety of proteomics services to assist your scientific research, including:

How to place an order

Please do not hesitate to reach out to us via email for a detailed discussion regarding your specific requirements. Our experienced experts will provide a feasible experimental scheme tailored to your specific requirements, ensuring high-quality results for protein analysis. Our customer service representatives are available 24 hours a day, from Monday to Sunday.

How to place an order

Reference

  1. Pandeswari PB, Sabareesh V. Middle-down approach: a choice to sequence and characterize proteins/proteomes by mass spectrometry. RSC Advances. 2019 Jan 2;9(1):313-344.

Q: How should gel band samples be collected and submitted for identification?

A: Sampling: Submit gel dots or gel bands stained with Coomassie Brilliant Blue or silver staining, ensuring clear and non-degraded bands.

Notes: a) Coomassie Brilliant Blue staining enhances the likelihood of identifying the target protein compared to silver staining. b) For selective identification of bands of interest, cut the specific bands and place them in an Eppendorf tube. c) If using gel strips, cut the entire lane and place it in an Eppendorf tube.

Sample Submission: After cutting the desired bands, add a few drops of double-distilled water to cover the bands, pack with ice packs, and send under chilled conditions (4°C).

Q: What are the precautions in the sampling process? Which components are not compatible with mass spectrometry?

A: In proteomics, proteasome inhibitors are not recommended during sample collection; also, try to avoid using solvents or extractants containing NP-40, Triton X-100, Tween 20, 80, high concentrations of SDS, etc. when preparing samples; Trypsin digestion is not recommended for adherent cell collection.

Q: What factors may contribute to protein degradation?

A: a) Inadequate handling during sample collection, such as prolonged collection time, introduction of contaminants (e.g., fermentation broth), failure of plant roots and leaves to absorb excess water after washing, elevated processing temperature, etc.;

b) Extended sample preparation duration leading to degradation;

c) Repetitive freeze-thaw cycles for the samples.

How to remove the interference of high abundance protein?

A: a) High-abundance proteins are mainly removed using high-abundance protein removal kits, including species-specific human, rat and mouse kits;

b) Use low abundance protein enrichment kits without species bias to enrich low abundance proteins;

c) For cases where there is no suitable product for removing high abundance proteins, SDS-PAGE cleavage and chromatographic separation can be used to achieve the goal of eliminating the interference of high abundance proteins.

Q: How to remove interference from high-abundance proteins?

A: a) Primarily utilize high-abundance protein depletion kits, including species-specific kits for humans, mice, etc., to remove high-abundance proteins. b) Employ species-neutral low-abundance protein enrichment kits to enrich low-abundance proteins. c) In cases where suitable high-abundance protein removal products are unavailable, methods such as SDS-PAGE gel cutting or chromatographic separation can be employed to eliminate interference from high-abundance proteins.

Q: In which samples are high-abundance proteins typically present?

A: a) Generally, bodily fluid samples such as urine, blood, milk, etc., often contain high-abundance proteins. b) In immunoprecipitation (IP) solution samples, high-abundance proteins may be present due to potential interference from antibodies.

Q: Which mass spectrometry platforms do you use for proteomics analysis?

A: We utilize the Thermo Orbitrap Fusion Lumos, Thermo Q Exactive series, Thermo Exploris 480, and timsTOF Pro for proteomics analysis.

Q: How can you determine if the extracted proteins are suitable for subsequent mass spectrometry analysis?

A: a) Clear and evenly distributed protein bands on SDS-PAGE; b) Good parallelism within sample replicates; c) Estimate the total extracted protein amount based on SDS-PAGE results. Generally, for regular proteomics, the protein total should be above 200 μg, and for modified proteomics, the total amount may need to be increased depending on the specific modification type; d) The protein concentration of the extracted sample should not be too low.

Q: Can enzymes related to apoptosis be detected using proteomics?

A: Proteomic analysis results include KEGG and GO annotations, allowing for the examination of protein annotations related to this function.

* For Research Use Only. Not for use in diagnostic procedures.
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