{"id":22995,"date":"2025-09-04T11:51:46","date_gmt":"2025-09-04T11:51:46","guid":{"rendered":"https:\/\/www.embl.org\/groups\/proteomics\/?page_id=22995"},"modified":"2026-04-23T08:54:05","modified_gmt":"2026-04-23T08:54:05","slug":"faqs","status":"publish","type":"page","link":"https:\/\/www.embl.org\/groups\/proteomics\/faqs\/","title":{"rendered":"FAQs"},"content":{"rendered":"\n<div class=\"vf-grid | vf-grid__col-1\"><div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<h2 class=\"wp-block-heading\">Frequently asked questions<\/h2>\n\n<\/div>\n<\/div>\n<\/div>\n\n\n\n<div class=\"vf-grid | vf-grid__col-5\"><div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<h3 class=\"wp-block-heading\">General information<\/h3>\n\n<\/div>\n<\/div>\n\n\n<div class=\"vf-grid__col--span-3\"><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nWho can use our services?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>Our services are primarily available to EMBL researchers. We also support external academic institutions. Services for industrial partners may be offered depending on our available capacity.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"contact-us-early\"  >\n<summary class=\"vf-details--summary\">\nContact us early to ensure the best results<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>To get the most out of your proteomics experiment, we strongly recommend reaching out to us as early as possible. Early contact allows us to support you in shaping the experimental design and ensuring optimal results.<\/p>\n\n\n\n<p>We can assist with key aspects such as:<\/p>\n\n\n\n<ul class=\"wp-block-list\">\n<li>Experimental design<br><\/li>\n\n\n\n<li>Determining the number of biological replicates<br><\/li>\n\n\n\n<li>Selecting appropriate controls<br><\/li>\n\n\n\n<li>Sample requirements (e.g., buffer conditions, sample volume)<\/li>\n<\/ul>\n\n\n\n<p><strong>To get started, please email us at <\/strong><a href=\"mailto:pcf@embl.de\"><strong>pcf@embl.de<\/strong><\/a><strong>.<\/strong><strong><br><\/strong>Include the following information to help us assess your project and guide you effectively:<\/p>\n\n\n\n<ul class=\"wp-block-list\">\n<li>Please provide your <strong>full name, address and group leader affiliation<\/strong>.<\/li>\n\n\n\n<li><strong>Type of analysis<\/strong> (e.g., full proteome analysis, immunoprecipitations, proximity labelling, phosphoproteomics, protein identification from Coomassie-stained gels, secretome analysis).<\/li>\n\n\n\n<li><strong>Sample type<\/strong> (e.g., cell line, primary cells, tissue, organs, animals)<\/li>\n\n\n\n<li><strong>Availability of a FASTA database<\/strong> for your species:<br>Databases can be found at:<a href=\"https:\/\/www.uniprot.org\/proteomes\"> https:\/\/www.uniprot.org\/proteomes<\/a><br>If you require a <strong>specific or custom database<\/strong>, please inform us in advance and provide the corresponding database for use in the analysis.<\/li>\n\n\n\n<li><strong>Number and annotation of samples <\/strong>(e.g., wild-type, mutant, domain-only, treatments, time course) <strong>and number of biological replicates<\/strong> (a minimum of 3 biological replicates is required for statistical analysis)<\/li>\n<\/ul>\n\n\n\n<p>By sharing this information, we can ensure that we design the most effective experiment tailored to your needs. This will also allow us to provide you with a rough cost estimate.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nWhat is the Service Request and why do I need it?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>We can only accept samples accompanied by a completed <a href=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2026\/01\/ServiceRequestForm_Externals.docx\" data-type=\"link\" data-id=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/08\/ServiceRequestForm_Externals.docx\">service request form<\/a>. This form provides all the essential information needed to process and analyze your samples.<br>A separate service request must be submitted for each individual set of samples.<\/p>\n\n\n\n<p>Please note:<\/p>\n\n\n\n<ul class=\"wp-block-list\">\n<li>Some sections of the form are only relevant to specific types of analysis. For example, gel images with annotated bands are not required for full proteome analysis.<\/li>\n\n\n\n<li>Protein identification relies on entries in a FASTA database. Overexpressed proteins from a different species will <strong>only<\/strong> be identified if they are included in the database.<\/li>\n\n\n\n<li>By default, we use the reference proteomes provided by UniProt for the respective organism when analyzing your samples. <br>If you require a <strong>specific or custom database<\/strong>, please inform us in advance and provide the corresponding database for use in the analysis.<\/li>\n<\/ul>\n\n\n\n<p>If you are analyzing a custom or modified protein, please provide its <strong>full amino acid sequence<\/strong>, including:<br><\/p>\n\n\n\n<ul class=\"wp-block-list\">\n<li>Linkers<br><\/li>\n\n\n\n<li>Cleavage sites<br><\/li>\n\n\n\n<li>Tags<br><\/li>\n\n\n\n<li>Any unnatural amino acids<\/li>\n<\/ul>\n\n\n\n<p>To help us interpret the sequence, please <strong>highlight tags, mutations, and cleavage sites in color<\/strong>.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\n[For internals only] Should I use the PPMS?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>EMBL Internal users have to submit their service request form via the PPMS system.&nbsp;<\/p>\n\n\n\n<p>For this an account in the system is necessary.&nbsp;Please follow this <a href=\"https:\/\/ppms.embl.de\/\" data-type=\"link\" data-id=\"https:\/\/ppms.embl.de\/\">link<\/a> to access the system.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nCan I submit instrument requests?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>Please note that you cannot request a certain instrument and we do not allow pre-booking of instrument time.<\/p>\n\n<\/div>\n<\/details>\n\n<\/div>\n<\/div>\n\n\n<div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<\/div>\n<\/div>\n<\/div>\n\n\n\n<hr class=\"vf-divider\">\n\n\n\n<div class=\"vf-grid | vf-grid__col-5\"><div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<h3 class=\"wp-block-heading\" id=\"info-submission\">Sample submission<\/h3>\n\n<\/div>\n<\/div>\n\n\n<div class=\"vf-grid__col--span-3\"><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nWhat are the general requirements?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>These are the essential requirements for standard procedures:<\/p>\n\n\n\n<figure class=\"wp-block-table is-style-regular\"><table class=\"has-fixed-layout\"><tbody><tr><td><strong>Experiment type<\/strong><\/td><td><strong>Material submitted to PCF<\/strong><\/td><\/tr><tr><td>Cell lysates<\/td><td>20 \u00b5g<\/td><\/tr><tr><td>Secretome, extracellular vesicles (EVs)<\/td><td>5-10 \u00b5g<\/td><\/tr><tr><td>Phosphoproteomics from lysates<\/td><td>500-1000 \u00b5g<\/td><\/tr><tr><td>Crosslinked protein complex<\/td><td>50 \u00b5g<\/td><\/tr><tr><td>Pull-downs, Immuno-precipitations<\/td><td>60 \u00b5L of eluate<\/td><\/tr><\/tbody><\/table><\/figure>\n\n\n\n<p><\/p>\n\n\n\n<p class=\"has-text-align-left\">Contaminants such as abundant serum proteins &#8211; especially from residual fetal calf serum (FCS) or bovine serum albumin (BSA) &#8211; can interfere with mass spectrometric analysis. To minimize these effects, we strongly recommend washing cultured cells three times with buffer such as PBS prior to lysis.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"compatible-buffers\"  >\n<summary class=\"vf-details--summary\">\nWhich buffers are compatible?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>Most commonly used lysis buffers are compatible with our protein digestion workflow, including:<\/p>\n\n\n\n<ul class=\"wp-block-list\">\n<li>Laemmli buffer (with or without bromophenol blue, \u03b2-mercaptoethanol, or dithiothreitol)<\/li>\n\n\n\n<li>SDS sample buffer<\/li>\n\n\n\n<li>RIPA buffer<\/li>\n\n\n\n<li>Others<\/li>\n<\/ul>\n\n\n\n<p>Our in-solution tryptic digestion is based on a modified SP3 protocol (<a href=\"https:\/\/doi.org\/10.1038\/s41596-018-0082-x\" data-type=\"link\" data-id=\"https:\/\/doi.org\/10.1038\/s41596-018-0082-x\" target=\"_blank\" rel=\"noreferrer noopener\">Hughes et al., 2019<\/a>, PMID: 30464214). For a detailed list of tested and compatible reagents, please refer to <strong>Table 2<\/strong> in the referenced publication.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nWhich additives are compatible?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>You may include <strong>EDTA-free protease inhibitor cocktails<\/strong> or <strong>phosphatase inhibitors<\/strong> in your lysis buffers, as they are compatible with our workflow.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nCan I submit viscous samples?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>If cell lysis results in the release of <strong>genomic DNA<\/strong>, the sample may become highly viscous. To reduce viscosity and ensure proper processing, please treat samples with <strong>benzonase<\/strong> or perform <strong>sonication<\/strong>. This step is also important for efficient recovery of <strong>nucleic acid-bound proteins<\/strong>.<\/p>\n\n<\/div>\n<\/details>\n\n<\/div>\n<\/div>\n\n\n<div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<\/div>\n<\/div>\n<\/div>\n\n\n\n<hr class=\"vf-divider\">\n\n\n\n<div class=\"vf-grid | vf-grid__col-5\"><div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<h3 class=\"wp-block-heading\">Protein Determination<\/h3>\n\n<\/div>\n<\/div>\n\n\n<div class=\"vf-grid__col--span-3\"><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nWhy do I need to determine my protein content before sample submission?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>Accurate protein quantification is essential for reliable <strong>mass spectrometry-based relative quantification<\/strong>. To ensure consistency and comparability across your samples, we require:<br><\/p>\n\n\n\n<ul class=\"wp-block-list\">\n<li><strong>Equal protein input<\/strong> for each condition and biological replicate<br><\/li>\n\n\n\n<li><strong>Accurate protein concentration measurement<\/strong>, ideally using a BSA standard curve to allow both relative and absolute quantification<\/li>\n<\/ul>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nWhat are the recommended methods?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>To determine protein concentration reliably, we recommend the following approaches:<br><\/p>\n\n\n\n<ul class=\"wp-block-list\">\n<li><strong>Tryptophan Assay<\/strong><strong><br><\/strong>Based on the method described by Wi\u015bniewski and Gaugaz (<a href=\"https:\/\/pubs.acs.org\/doi\/10.1021\/ac504689z\">DOI: 10.1021\/ac504689z<\/a>), this fluorescence-based assay is cost-effective, user-friendly, and compatible with a wide range of sample types and buffer components.<br><em>Note: This assay requires black microplates and a fluorescence-capable plate reader. Please check compatibility with your equipment.<\/em><em><br><\/em><\/li>\n\n\n\n<li><strong>Colorimetric Assays<\/strong>\n<ul class=\"wp-block-list\">\n<li><strong>Bradford assay<\/strong><\/li>\n\n\n\n<li><strong>BCA (Bicinchoninic Acid) assay<\/strong><br>These are widely used and generally reliable for protein concentration determination.<\/li>\n<\/ul>\n<\/li>\n<\/ul>\n\n\n\n<p><strong>Important Note<br><\/strong>Protein concentration measurements using <strong>NanoDrop or similar spectrophotometers<\/strong> are <strong>not reliable<\/strong> and should be avoided.<\/p>\n\n<\/div>\n<\/details>\n\n<\/div>\n<\/div>\n\n\n<div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<\/div>\n<\/div>\n<\/div>\n\n\n\n<hr class=\"vf-divider\">\n\n\n\n<div class=\"vf-grid | vf-grid__col-5\"><div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<h3 class=\"wp-block-heading\" id=\"info-sample-specific\">Information for specific sample types<\/h3>\n\n<\/div>\n<\/div>\n\n\n<div class=\"vf-grid__col--span-3\"><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nGeneral information<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>In general, we recommend performing all possible quality control steps to ensure the success of the mass spectrometry analysis of your samples. These steps may include protein quantification, SDS-PAGE combined with Coomassie or silver staining, and Western blot analyses to verify knockout, expression, or enrichment of the protein of interest. However, we understand that sometimes it is necessary to proceed with mass spectrometry analysis without controlling every step, for example, when working with limited sample material.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nCan I submit samples at peptide-level?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>If you plan to submit samples at the <strong>peptide level<\/strong>, please <a href=\"mailto:pcf@embl.de\" data-type=\"mailto\" data-id=\"mailto:pcf@embl.de\">contact us<\/a> <strong>in advance<\/strong>. Note that we can only accept peptide samples prepared in <strong>mass spectrometry-compatible buffers<\/strong>.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nHow do I submit full proteome samples?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>For a full proteome analysis, we recommend using buffers with harsh detergents such as SDS to ensure complete cell lysis. A suitable choice is RIPA buffer (0.1% SDS, 1% deoxycholate, 1% NP-40, 150 mM NaCl in 50 mM Tris\/HCl, pH 7\u20138), supplemented with EDTA-free protease inhibitors. To reduce sample viscosity and facilitate protein extraction, it is essential to degrade DNA by adding Benzonase or using sonication, followed by removal of cell debris via centrifugation.<br>Accurate protein quantification is critical to provide equal protein amounts for reliable quantitative comparison between different conditions. We recommend determining protein concentration using BCA, Bradford, or Tryptophan assays, as methods like NanoDrop are not sufficiently accurate. Please adjust all samples to 20 \u00b5g of protein in 60 \u00b5L of lysis buffer (e.g., Laemmli buffer). Insufficient protein amounts may lead to incomplete proteomic analysis, while excessive protein concentrations can reduce digestion and TMT labeling efficiency.<br><br>If you cannot provide 20 \u00b5g per sample, please contact us to discuss alternatives.<br>For further details, please also see the <a href=\"#submission-info\" data-type=\"internal\" data-id=\"#faq-samplesubmission\">Sample Submission section<\/a>.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nHow do I submit immuno-precipitation (IP), pull-down (PD) or proximity labeling samples?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<ul class=\"wp-block-list\">\n<li>Please <strong>do not perform protein quantification<\/strong> on eluates from IP or proximity labeling experiments. Instead, submit <strong>equal elution volumes<\/strong> for all samples, providing no more than <strong>60 \u00b5L of eluate per sample<\/strong>.<br><\/li>\n\n\n\n<li>Ensure that <strong>all beads are completely removed<\/strong> before submission.<br><\/li>\n\n\n\n<li>Use an <strong>equal number of cells<\/strong> or equivalent protein amounts as starting material for each sample.<\/li>\n<\/ul>\n\n\n\n<p>To isolate protein complexes, buffers containing <strong>0.1% Triton X-100 with 150 mM NaCl in 100 mM HEPES\/NaOH (pH ~7.5)<\/strong> are commonly used as a good starting point. However, we recommend optimizing lysis conditions for your specific protein of interest. More detailed guidance on lysis and immunoaffinity purification can be found <a href=\"https:\/\/www.thermofisher.com\/de\/de\/home\/life-science\/protein-biology\/protein-biology-learning-center\/protein-biology-resource-library\/pierce-protein-methods\/immunoprecipitation-ip.html\" data-type=\"link\" data-id=\"https:\/\/www.thermofisher.com\/de\/de\/home\/life-science\/protein-biology\/protein-biology-learning-center\/protein-biology-resource-library\/pierce-protein-methods\/immunoprecipitation-ip.html\">here<\/a>.<\/p>\n\n\n\n<h4 class=\"wp-block-heading\">Special considerations<\/h4>\n\n\n\n<ul class=\"wp-block-list\">\n<li>Immunoaffinity purification of <strong>nuclear proteins<\/strong> can be challenging due to the harsh lysis conditions required. For enrichment of nuclear protein complexes, we recommend <strong>proximity labeling<\/strong> approaches.<br><\/li>\n\n\n\n<li>Please note that <strong>sonication<\/strong> may affect the integrity of protein complexes and should be used with caution.<\/li>\n<\/ul>\n\n\n\n<h4 class=\"wp-block-heading\"><strong>Quality Control Recommendations<\/strong><\/h4>\n\n\n\n<ul class=\"wp-block-list\">\n<li>Perform a <strong>Western blot<\/strong> to verify enrichment of your protein of interest and to check for co-immunoprecipitation of known interaction partners.<br><\/li>\n\n\n\n<li>Run <strong>SDS-PAGE<\/strong> followed by <strong>Coomassie or silver staining<\/strong> depending on sample amount. Confirm that control samples contain proteins and that the bait-IP sample shows reduced complexity compared to the input lysate.<br><\/li>\n\n\n\n<li><strong>Elution with Laemmli buffer<\/strong> is generally effective for releasing protein complexes.<br><\/li>\n\n\n\n<li>We <strong>do not recommend<\/strong> using <strong>formaldehyde-based crosslinking<\/strong>.<\/li>\n<\/ul>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nHow do I submit secretome samples?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>For secretome analysis, it is essential to culture cells in <strong>serum-free medium<\/strong>. The presence of abundant proteins, such as those from fetal calf\/bovine serum, can dominate the sample and produce prominent peptides. Since the mass spectrometer prioritizes analyzing the most abundant peptides first, this can lead to detecting serum proteins like albumin instead of the proteins of interest in your sample.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nHow do I submit extracellular matrix samples?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>We provide services for the mass spectrometric analysis of extracellular matrices (ECMs). Below is a selection of key publications relevant to ECM proteomics and sample preparation:<\/p>\n\n\n\n<ul class=\"wp-block-list\">\n<li>Khan MM, Galea G, Jung J, et al. <em>Dextromethorphan inhibits collagen and collagen-like cargo secretion to ameliorate lung fibrosis.<\/em> Science Translational Medicine. 2024 Dec;16(778):eadj3087. DOI: <a href=\"https:\/\/doi.org\/10.1126\/scitranslmed.adj3087\" data-type=\"link\" data-id=\"https:\/\/doi.org\/10.1126\/scitranslmed.adj3087\">10.1126\/scitranslmed.adj3087<\/a>. PMID: 39693409.<br><\/li>\n\n\n\n<li>Naba A, Clauser KR, Hynes RO. <em>Enrichment of Extracellular Matrix Proteins from Tissues and Digestion into Peptides for Mass Spectrometry Analysis.<\/em> J Vis Exp. 2015;(101):e53057. DOI: <a href=\"https:\/\/doi.org\/10.3791\/53057\" data-type=\"link\" data-id=\"https:\/\/doi.org\/10.3791\/53057\">10.3791\/53057<\/a>. PMID: 26273955.<br><\/li>\n\n\n\n<li>McCabe MC, Schmitt LR, Hill RC, et al. <em>Evaluation and Refinement of Sample Preparation Methods for Extracellular Matrix Proteome Coverage.<\/em> Mol Cell Proteomics. 2021 Jan;20:100079. DOI: <a href=\"https:\/\/doi.org\/10.1016\/j.mcpro.2021.100079\" data-type=\"link\" data-id=\"https:\/\/doi.org\/10.1016\/j.mcpro.2021.100079\">10.1016\/j.mcpro.2021.100079<\/a>. PMID: 33845168.<\/li>\n<\/ul>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nHow do I submit samples for phosphoproteomic analysis<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>Phosphoproteomic analysis requires a <strong>high amount of input material<\/strong>\u2014typically around <strong>500 \u00b5g of total protein<\/strong>. The number of phosphopeptides detected is strongly dependent on the amount of starting material provided.<\/p>\n\n\n\n<p>To ensure optimal results, please <strong><a href=\"mailto:pcf@embl.de\" data-type=\"mailto\" data-id=\"mailto:pcf@embl.de\">contact us<\/a> in advance<\/strong> to receive our <strong>lysis protocol<\/strong>, which is specifically optimized for compatibility with our downstream phosphopeptide enrichment workflow.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nHow do I submit samples for plasma or serum analysis<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>Plasma and serum contain a few very high-abundance proteins (e.g., albumin, immunoglobulins) that can interfere with the in-depth analysis of lower-abundance proteins by mass spectrometry.<\/p>\n\n\n\n<p>To improve proteome coverage, we recommend using the ENRICH-iST kit from PreOmics, which is specifically designed for the depletion of abundant plasma proteins:<a href=\"https:\/\/www.preomics.com\/products\/enrich-ist\"> https:\/\/www.preomics.com\/products\/enrich-ist<\/a><\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nHow do I submit samples for in-gel analysis<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<ul class=\"wp-block-list\">\n<li>Please send <strong>only cut gel bands<\/strong> from <strong>Coomassie-stained gels<\/strong>.<br><\/li>\n\n\n\n<li>Ensure your Coomassie stain is <strong>MS-compatible<\/strong> by consulting the manufacturer\u2019s datasheet or contacting their customer service.<br><\/li>\n\n\n\n<li>Follow the instructions outlined in our <strong><a href=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/10\/Excision-of-gel-bands.pdf\" data-type=\"page\" data-id=\"96\" target=\"_blank\" rel=\"noreferrer noopener\">gel-cutting procedure<\/a><\/strong>.<br><\/li>\n\n\n\n<li><strong>Gel images are mandatory<\/strong>; samples without an accompanying gel image will not be accepted.<br><\/li>\n\n\n\n<li>Clearly <strong>label the protein ladder<\/strong> and number each cut band.<br><\/li>\n\n\n\n<li>Mark the exact area excised from the gel.<br><\/li>\n\n\n\n<li>Include <strong>empty lanes between samples<\/strong> and avoid overloading.<br><\/li>\n\n\n\n<li>Note that the <strong>quality of your gel directly affects the accuracy<\/strong> of MS analysis results.<\/li>\n<\/ul>\n\n\n\n<p><\/p>\n\n<\/div>\n<\/details>\n\n<\/div>\n<\/div>\n\n\n<div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<\/div>\n<\/div>\n<\/div>\n\n\n\n<hr class=\"vf-divider\">\n\n\n\n<div class=\"vf-grid | vf-grid__col-5\"><div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<h3 class=\"wp-block-heading\">Sample preparation<\/h3>\n\n<\/div>\n<\/div>\n\n\n<div class=\"vf-grid__col--span-3\"><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<details  class=\"vf-details\" id=\"sample-prep-guidelines\"  >\n<summary class=\"vf-details--summary\">\nHow do I best prepare my samples?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<ul class=\"wp-block-list\">\n<li>Wear <strong>gloves <\/strong>at all times\n<ul class=\"wp-block-list\">\n<li>Always wear gloves during sample preparation to prevent introducing contaminants, such as human keratins, into your samples. Ensure gloves are changed regularly.<\/li>\n<\/ul>\n<\/li>\n\n\n\n<li>Use <strong>proper <\/strong>sample <strong>handling and storage<\/strong> conditions\n<ul class=\"wp-block-list\">\n<li>Store samples in conditions that preserve their integrity for mass spectrometric analysis. If freezing is required, use appropriate vials and store at -80\u00b0C or lower, depending on your sample\u2019s needs.<\/li>\n<\/ul>\n<\/li>\n\n\n\n<li><strong>Label <\/strong>samples <strong>clearly<\/strong>\n<ul class=\"wp-block-list\">\n<li>Ensure that all samples are clearly labeled with your initials and single consecutive numbers (avoid detailed explanations on the tubes). Detailed explanations should be provided in our service request form.<\/li>\n<\/ul>\n<\/li>\n<\/ul>\n\n\n\n<ul class=\"wp-block-list\">\n<li>Use <strong>fresh reagents<\/strong> for buffer preparation\n<ul class=\"wp-block-list\">\n<li>Always prepare buffers using fresh stocks and chemicals. Older chemicals may be a source of cross-contamination or keratin contamination. Additionally, some chemicals degrade, oxidize, or attract moisture over time, which can affect the quality and consistency of your samples.<\/li>\n<\/ul>\n<\/li>\n\n\n\n<li><strong>Clean <\/strong>equipment regularly\n<ul class=\"wp-block-list\">\n<li>Clean all tools, surfaces, and equipment thoroughly with 70% ethanol before and after use. Use lint-free wipes (e.g., Kimberly-Clark Kimtech Science) to ensure no fibers or debris are left behind.<\/li>\n<\/ul>\n<\/li>\n<\/ul>\n\n\n\n<div style=\"height:40px\" aria-hidden=\"true\" class=\"wp-block-spacer\"><\/div>\n\n\n\n<p><span style=\"text-decoration: underline;\">For gel samples only<\/span>:<\/p>\n\n\n\n<ul class=\"wp-block-list\">\n<li>Use <strong>clean containers<\/strong> for gels<\/li>\n\n\n\n<li><strong>Cover <\/strong>with a lid\n<ul class=\"wp-block-list\">\n<li>Ensure that containers used for storing gels are always sealed with a lid to prevent contamination and exposure to airborne particles.<\/li>\n<\/ul>\n<\/li>\n\n\n\n<li>Prepare gel bands on <strong>clean surfaces<\/strong><\/li>\n<\/ul>\n\n<\/div>\n<\/details>\n\n<\/div>\n<\/div>\n\n\n<div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<\/div>\n<\/div>\n<\/div>\n\n\n\n<hr class=\"vf-divider\">\n\n\n\n<div class=\"vf-grid | vf-grid__col-5\"><div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<h3 class=\"wp-block-heading\" id=\"submission-info\">Submission information<\/h3>\n\n<\/div>\n<\/div>\n\n\n<div class=\"vf-grid__col--span-3\"><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nWhat is important when I submit samples?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<ul class=\"wp-block-list\">\n<li><strong><a href=\"mailto:pcf@embl.de\" data-type=\"mailto\" data-id=\"mailto:pcf@embl.de\">Contact Us<\/a> First:<\/strong> Do not send samples without prior discussion with PCF staff.<br><\/li>\n\n\n\n<li><strong>Packaging:<\/strong> Use secure, leak-proof tubes. <em>Do not use parafilm.<\/em> Clearly label each tube with your initials and a sequential number.<br><\/li>\n\n\n\n<li><strong>Sample Preparation:<\/strong> Spin down and freeze all protein samples. Place all tubes into a <strong>single container<\/strong> (e.g., a box or bag) <strong>inside the styrofoam shipping box<\/strong>. This ensures all samples remain together and protected during transit.<br><\/li>\n\n\n\n<li><strong>Documentation:<\/strong> Include a printed copy of the completed <a href=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2026\/01\/ServiceRequestForm_Externals.docx\" data-type=\"link\" data-id=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/08\/ServiceRequestForm_Externals.docx\">service request form<\/a> in a transparent sleeve inside the shipment. <em>(Example image)<\/em><br>The form must list all treatments applied to each sample, in the same order as the samples are arranged in the box.<\/li>\n<\/ul>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nHow do I ship my samples?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<ul class=\"wp-block-list\">\n<li><strong>Ship <\/strong>to the following address:<\/li>\n<\/ul>\n\n\n\n<div class=\"vf-box vf-box--very-easy | vf-u-margin__bottom--400\">\n      <h3 class=\"vf-box__heading\">\n                Shipping address                  <\/h3> \n        <p style=\"text-align: left;\"><strong>EMBL Heidelberg<\/strong><br \/>\n*your contact person at PCF*<br \/>\nProteomics Core Facility<br \/>\nMeyerhofstr. 1<br \/>\n69117 Heidelberg<br \/>\nGermany<\/p>\n<\/div>\n\n\n<ul class=\"wp-block-list\">\n<li><strong>Timing:<\/strong> Avoid shipping during German public holidays. Ship early in the week to ensure samples arrive before the weekend. Do not ship over the weekend.<br><\/li>\n\n\n\n<li><strong>Notification:<\/strong> Inform PCF staff once samples are sent. Email a digital copy of the <a href=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2026\/01\/ServiceRequestForm_Externals.docx\" data-type=\"link\" data-id=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/08\/ServiceRequestForm_Externals.docx\">service request<\/a> form to <a href=\"mailto:pcf@embl.de\">pcf@embl.de<\/a>.<\/li>\n<\/ul>\n\n\n\n<h4 class=\"wp-block-heading\"><strong>Additional Information for shipments from non-EU countries<\/strong><\/h4>\n\n\n\n<p>In order to avoid any delay in receiving your samples, please follow our <a href=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/08\/Shipping_Guidelines_2025.pdf\" data-type=\"link\" data-id=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/08\/Shipping_Guidelines_2025.pdf\">requirements for shipping samples<\/a> from non-EU countries and add a <a href=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/08\/Proforma_Invoice_Templates.docx\" data-type=\"link\" data-id=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/08\/Proforma_Invoice_Templates.docx\">proforma invoice<\/a> to your shipment.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nHow long will sample processing take?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<ul class=\"wp-block-list\">\n<li><strong>In-gel samples<\/strong> are processed weekly, typically on <strong>Thursdays<\/strong>. Gels received by <strong>Wednesday evening<\/strong> will be included in that week\u2019s run. If no technical issues arise, results can usually be expected the <strong>following week<\/strong>.<\/li>\n\n\n\n<li><strong>In-solution samples<\/strong> have a standard turnaround time of <strong>4 to 6 weeks<\/strong> from the date of receipt.<\/li>\n<\/ul>\n\n\n\n<p><strong>Please note:<\/strong> During periods of high demand, processing times may be longer than stated.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nHow long are my samples and data stored?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>Please note that upon completion of the project (after receipt of the data) we will store the remaining <span style=\"text-decoration: underline;\">samples <\/span>for <strong>8 weeks<\/strong>.<\/p>\n\n\n\n<p>Please also note that we do not send any samples back.<\/p>\n\n\n\n<p>The <span style=\"text-decoration: underline;\">raw files<\/span> of your experiment are archived for <strong>10 years<\/strong> on our server.<\/p>\n\n<\/div>\n<\/details>\n\n<\/div>\n<\/div>\n\n\n<div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<\/div>\n<\/div>\n<\/div>\n\n\n\n<hr class=\"vf-divider\">\n\n\n\n<div class=\"vf-grid | vf-grid__col-5\"><div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<h3 class=\"wp-block-heading\">Prices<\/h3>\n\n<\/div>\n<\/div>\n\n\n<div class=\"vf-grid__col--span-3\"><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nHow much does it cost to analyse my samples?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<ul class=\"wp-block-list\">\n<li><strong>Internal rates<\/strong> apply to current <strong>EMBL members<\/strong> and active <a href=\"https:\/\/yip-search.embo.org\/#\/\">EMBO Young Investigator \/ EMBO Installation Grantee \/ EMBO Global Investigator.<\/a><\/li>\n<\/ul>\n\n\n\n<ul class=\"wp-block-list\">\n<li><strong>External rates<\/strong> (as per our published price list) apply to <strong>academic institutions<\/strong> and <strong>former EMBL members<\/strong>.<\/li>\n<\/ul>\n\n\n\n<ul class=\"wp-block-list\">\n<li>We do <strong>not offer discounts<\/strong> for higher sample numbers.<br><\/li>\n\n\n\n<li>Projects are generally <strong>not offered on a collaborative basis only<\/strong>, as cost recovery for consumables and instrument maintenance is required.<br><strong>Please note:<\/strong> All experiments, including <strong>pilot studies<\/strong>, must be fully compensated.<br><\/li>\n\n\n\n<li><strong>Industry partners<\/strong> are subject to <strong>special pricing<\/strong>.<\/li>\n<\/ul>\n\n\n\n<p><strong>Please note:<\/strong> Our facility is charging solely for the consumables and measurement time. Costs depend on the sample type, complexity and number of samples.<\/p>\n\n\n\n<p>These are our current fees for external customers (as of August 2025):<\/p>\n\n\n\n<figure class=\"wp-block-table alignleft\"><table><thead><tr><th><\/th><th><strong>Prices for <\/strong><br><strong>external customers<\/strong><\/th><\/tr><\/thead><tbody><tr><td>Sample processing fee (per sample)<\/td><td>3 \u20ac<\/td><\/tr><tr><td>TMT6-plex (up to 6 samples)<\/td><td>100 \u20ac<\/td><\/tr><tr><td>TMT11-plex (up to 11 samples)<\/td><td>200 \u20ac<\/td><\/tr><tr><td>TMT18-plex (up to 18 samples)<\/td><td>280 \u20ac<\/td><\/tr><tr><td>Sample desalting (per sample)<\/td><td>20 \u20ac<\/td><\/tr><tr><td>Phosphoenrichment incl. desalting (per sample)<\/td><td>20 \u20ac<\/td><\/tr><tr><td>Peptide-SEC \/ high pH \/ PGC fractionation (per sample)<\/td><td>20 \u20ac<\/td><\/tr><tr><td>Measurement time (per hour)<\/td><td>100 \u20ac<\/td><\/tr><\/tbody><\/table><figcaption class=\"wp-element-caption\">PCF service fees for the indicated items for our external customers (as of August 2025)<\/figcaption><\/figure>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nHow do you calculate prices?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<div class=\"vf-tabs\"><ul class=\"vf-tabs__list\" data-vf-js-tabs=\"true\"><li class=\"vf-tabs__item\"><a class=\"vf-tabs__link\" href=\"#vf-tabs__section-e230dbbe-f1c3-4956-81f3-a0e91e5f4682\" data-vf-js-location-nearest-activation-target=\"\">Example 1 | Full proteome with TMT<\/a><\/li><li class=\"vf-tabs__item\"><a class=\"vf-tabs__link\" href=\"#vf-tabs__section-de7656ab-cf26-4089-8c7c-25584f22eae2\" data-vf-js-location-nearest-activation-target=\"\">Example 2 | Full proteome with DIA<\/a><\/li><li class=\"vf-tabs__item\"><a class=\"vf-tabs__link\" href=\"#vf-tabs__section-f56481a7-f6ff-417f-8ea6-7bd1c02e94a3\" data-vf-js-location-nearest-activation-target=\"\">Example 3 | IP-like with TMT<\/a><\/li><li class=\"vf-tabs__item\"><a class=\"vf-tabs__link\" href=\"#vf-tabs__section-3736d0f9-10a2-4157-acb4-8053e6268664\" data-vf-js-location-nearest-activation-target=\"\">Example 4 | In-gel samples<\/a><\/li><\/ul><div class=\"vf-tabs-content\" data-vf-js-tabs-content=\"true\">\n<section class=\"vf-tabs__section\" id=\"vf-tabs__section-e230dbbe-f1c3-4956-81f3-a0e91e5f4682\"><h2>Example 1 | Full proteome with TMT<\/h2>\n<p>You have six conditions, each with three replicates, and you want to look at changes in protein expression on the whole proteome level (e.g. from cell lysates, tissue lysates, or subcellular fractions).<\/p>\n\n\n\n<p>This would be a TMT18 experiment (18 samples total), and the cost is \u20ac3,044 (excluding tax), which includes our standard data analysis.<\/p>\n\n\n\n<figure class=\"wp-block-table alignleft\"><table class=\"has-fixed-layout\"><thead><tr><th><\/th><th>Price<\/th><\/tr><\/thead><tbody><tr><td>TMT18-plex labelling reagent<\/td><td>1x 280 \u20ac<\/td><\/tr><tr><td>high pH fractionation<\/td><td>1x 20 \u20ac<\/td><\/tr><tr><td>desalting<\/td><td>1x 20 \u20ac<\/td><\/tr><tr><td>consumables<\/td><td>54 \u20ac (18x 3 \u20ac)<\/td><\/tr><tr><td>pre-Run of 90 min + 30 min blank run<\/td><td>200 \u20ac (100 \u20ac\/h measurement time)<\/td><\/tr><tr><td>120 min measurement of 12 fractions, followed by 30 min blank run<\/td><td>2450 \u20ac (total of 24.5 h)<\/td><\/tr><\/tbody><tfoot><tr><td>Total sum (without VAT)<\/td><td>3044 \u20ac<\/td><\/tr><\/tfoot><\/table><figcaption class=\"wp-element-caption\">Example of price calculation for the analysis of a full proteome with a total of 18 samples in a TMT18-based experiment.<\/figcaption><\/figure>\n<\/section>\n\n\n\n<section class=\"vf-tabs__section\" id=\"vf-tabs__section-de7656ab-cf26-4089-8c7c-25584f22eae2\"><h2>Example 2 | Full proteome with DIA<\/h2>\n<p>You have 20 samples that you would like to measure in DIA mode (recommended for tissue samples, muscle cell lines, or in case you have more than 18 sample to compare)<\/p>\n\n\n\n<figure class=\"wp-block-table alignleft\"><table class=\"has-fixed-layout\"><thead><tr><th><\/th><th>Price<\/th><\/tr><\/thead><tbody><tr><td>consumables<\/td><td>60 \u20ac (20 x 3 \u20ac)<\/td><\/tr><tr><td>120 min measurement of 20 samples, followed by 30 min blank runs<\/td><td>5000 \u20ac (total of 50 h; 100 \u20ac\/h measurement time)<\/td><\/tr><\/tbody><tfoot><tr><td>Total sum (without VAT)<\/td><td>5060 \u20ac<\/td><\/tr><\/tfoot><\/table><figcaption class=\"wp-element-caption\">Example of price calculation for the analysis of a full proteome with a total of 20 samples in a DIA-based experiment.<\/figcaption><\/figure>\n<\/section>\n\n\n\n<section class=\"vf-tabs__section\" id=\"vf-tabs__section-f56481a7-f6ff-417f-8ea6-7bd1c02e94a3\"><h2>Example 3 | IP-like with TMT<\/h2>\n<p>You have a co-immunoprecipitation in triplicates (3x bait IP +&nbsp; 3x control IP). This would be a TMT6 approach. Since you purify a certain protein along with its interaction partners and some contaminants you expect a reduced set of proteins to be detected (\u201cmedium\u201d complexity sample) as compared to a full proteome sample (e.g lysates).<\/p>\n\n\n\n<figure class=\"wp-block-table alignleft\"><table class=\"has-fixed-layout\"><thead><tr><th><\/th><th>Price<\/th><\/tr><\/thead><tbody><tr><td>TMT6-plex labelling reagent<\/td><td>1x 100 \u20ac<\/td><\/tr><tr><td>high pH fractionation<\/td><td>1x 20 \u20ac<\/td><\/tr><tr><td>desalting<\/td><td>1x 20 \u20ac<\/td><\/tr><tr><td>consumables<\/td><td>18 \u20ac (6 x 3 \u20ac)<\/td><\/tr><tr><td>pre-Run of 60 min + 30 min blank run<\/td><td>150 \u20ac (100 \u20ac\/h measurement time)<\/td><\/tr><tr><td>60 min measurement of 6 fractions, followed by 30 min blank run<\/td><td>650 \u20ac (total of 6.5 h; 100 \u20ac\/h measurement time)<\/td><\/tr><\/tbody><tfoot><tr><td>Total sum (without VAT)<\/td><td>978 \u20ac<\/td><\/tr><\/tfoot><\/table><figcaption class=\"wp-element-caption\">Example of price calculation for the analysis of an IP-like sample with a total of 6 samples in a TMT6-based experiment.<\/figcaption><\/figure>\n<\/section>\n\n\n\n<section class=\"vf-tabs__section\" id=\"vf-tabs__section-3736d0f9-10a2-4157-acb4-8053e6268664\"><h2>Example 4 | In-gel samples<\/h2>\n<p>You have a prominent band on a Coomassie-stained gel and want to identify or confirm the protein. This is considered a low-complexity sample, since we expect only a few proteins to be present.<\/p>\n\n\n\n<p>The cost for analyzing each band is \u20ac103.<\/p>\n\n\n\n<figure class=\"wp-block-table alignleft\"><table class=\"has-fixed-layout\"><thead><tr><th><\/th><th>Price<\/th><\/tr><\/thead><tbody><tr><td>consumables<\/td><td>3\u20ac (1 x 3 \u20ac)<\/td><\/tr><tr><td>30 min measurement of sample, followed by 30 min blank run<\/td><td>100 \u20ac (total of 1 h; 100 \u20ac\/h measurement time)<\/td><\/tr><\/tbody><tfoot><tr><td>Total sum (without VAT)<\/td><td>103 \u20ac<\/td><\/tr><\/tfoot><\/table><figcaption class=\"wp-element-caption\">Example of price calculation for the analysis of one SDS-PAGE gel band for protein identification.<\/figcaption><\/figure>\n<\/section>\n<\/div><\/div>\n\n<\/div>\n<\/details>\n\n<\/div>\n<\/div>\n\n\n<div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<\/div>\n<\/div>\n<\/div>\n\n\n\n<hr class=\"vf-divider\">\n\n\n\n<div class=\"vf-grid | vf-grid__col-5\"><div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<h3 class=\"wp-block-heading\">Computational support &amp; results<\/h3>\n\n<\/div>\n<\/div>\n\n\n<div class=\"vf-grid__col--span-3\"><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nWhat will my results look like?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>We deliver a comprehensive data analysis of the proteomics experiment along with the complete R script to ensure full reproducibility. Our pipeline includes contaminant removal, correction for batch effects, variance-stabilising normalisation and &#8211; if required &#8211; data imputation. Differential expression is assessed using moderated t-statistics from the limma package, and results are visualised through correlation plots and heat maps with clustering to highlight patterns in the data. To support biological interpretation, we also provide a basic Gene Ontology enrichment analysis for standard model organisms.<\/p>\n\n\n\n<div class=\"vf-grid | vf-grid__col-4\"><div class=\"vf-grid__col--span-2\"><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<h5 class=\"wp-block-heading has-text-align-center\">\n  Data normalization\n<\/h5>\n\n\n\n<figure class=\"vf-figure wp-block-image  | vf-figure--align vf-figure--align-centered  size-large is-style-default wp-duotone-unset-1\"><img loading=\"lazy\" decoding=\"async\" width=\"1024\" height=\"717\" class=\"vf-figure__image\" src=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Normalization_overview_sub_V1-1024x717.png\" alt=\"\" class=\"wp-image-24683\" srcset=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Normalization_overview_sub_V1-1024x717.png 1024w, https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Normalization_overview_sub_V1-300x210.png 300w, https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Normalization_overview_sub_V1-768x538.png 768w, https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Normalization_overview_sub_V1.png 1500w\" sizes=\"auto, (max-width: 1024px) 100vw, 1024px\" \/><\/figure>\n\n<\/div>\n<\/div>\n\n\n<div class=\"vf-grid__col--span-2\"><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<h5 class=\"wp-block-heading has-text-align-center\">Principal component analysis (PCA)<\/h5>\n\n\n\n<figure class=\"vf-figure wp-block-image  | vf-figure--align vf-figure--align-centered  size-large\"><img loading=\"lazy\" decoding=\"async\" width=\"1024\" height=\"657\" class=\"vf-figure__image\" src=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/PCA_analysis_sub_V1-1-1024x657.png\" alt=\"\" class=\"wp-image-24703\" srcset=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/PCA_analysis_sub_V1-1-1024x657.png 1024w, https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/PCA_analysis_sub_V1-1-300x192.png 300w, https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/PCA_analysis_sub_V1-1-768x493.png 768w, https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/PCA_analysis_sub_V1-1.png 1500w\" sizes=\"auto, (max-width: 1024px) 100vw, 1024px\" \/><\/figure>\n\n<\/div>\n<\/div>\n<\/div>\n\n\n\n<div class=\"vf-grid | vf-grid__col-4\"><div class=\"vf-grid__col--span-2\"><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<h5 class=\"wp-block-heading has-text-align-center\">Volcano plot of differentially abundant proteins<\/h5>\n\n\n\n<figure class=\"vf-figure wp-block-image  | vf-figure--align vf-figure--align-centered  size-large\"><img loading=\"lazy\" decoding=\"async\" width=\"1024\" height=\"655\" class=\"vf-figure__image\" src=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Volcano_plot_small_V1-1-1024x655.png\" alt=\"\" class=\"wp-image-24705\" srcset=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Volcano_plot_small_V1-1-1024x655.png 1024w, https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Volcano_plot_small_V1-1-300x192.png 300w, https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Volcano_plot_small_V1-1-768x492.png 768w, https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Volcano_plot_small_V1-1.png 1500w\" sizes=\"auto, (max-width: 1024px) 100vw, 1024px\" \/><\/figure>\n\n<\/div>\n<\/div>\n\n\n<div class=\"vf-grid__col--span-2\"><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<h5 class=\"wp-block-heading has-text-align-center\">Cluster analysis<\/h5>\n\n\n\n<figure class=\"vf-figure wp-block-image  | vf-figure--align vf-figure--align-centered  size-large\"><img loading=\"lazy\" decoding=\"async\" width=\"1024\" height=\"731\" class=\"vf-figure__image\" src=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Heatmap_hits_sub_5_proteins_V1-1024x731.png\" alt=\"\" class=\"wp-image-24679\" srcset=\"https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Heatmap_hits_sub_5_proteins_V1-1024x731.png 1024w, https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Heatmap_hits_sub_5_proteins_V1-300x214.png 300w, https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Heatmap_hits_sub_5_proteins_V1-768x549.png 768w, https:\/\/www.embl.org\/groups\/proteomics\/wp-content\/uploads\/2025\/09\/Heatmap_hits_sub_5_proteins_V1.png 1050w\" sizes=\"auto, (max-width: 1024px) 100vw, 1024px\" \/><\/figure>\n\n<\/div>\n<\/div>\n<\/div>\n\n\n\n<p>For data analysis, we primarily use <a href=\"https:\/\/www.uniprot.org\/proteomes\" data-type=\"link\" data-id=\"https:\/\/www.uniprot.org\/proteomes\">UniProt Proteome Databases<\/a>, which include only one representative entry per gene. As a result, information on individual isoforms is not reported.<br>If you require a <strong>specific or custom database<\/strong>, please inform us in advance and provide the corresponding database for use in the analysis.<\/p>\n\n\n\n<p>Please consider whether this level of analysis is suitable for your research question. If distinguishing between isoforms is important for your study, let us know in advance so we can discuss possible options.<\/p>\n\n<\/div>\n<\/details>\n\n\n\n<details  class=\"vf-details\" id=\"\"  >\n<summary class=\"vf-details--summary\">\nWhat can I expect from a typical proteome sample?<\/summary>\n<div class=\"acf-innerblocks-container\">\n\n<p>A <strong>full proteome<\/strong> analysis of an immortalized cell line, such as HEK293T cells, typically yields around 8,000 identified protein groups (with a minimum of 1 peptide per protein), of which approximately 7,000 proteins are quantified (minimum 2 peptides per protein) across compared conditions.<\/p>\n\n\n\n<p>For other cell types, such as <em>Escherichia coli<\/em>, we generally identify about 2,200 proteins, with around 2,000 proteins quantified. These numbers vary significantly depending on sample type and proteome complexity.<\/p>\n\n\n\n<p>For samples like <strong>tissue lysates<\/strong>, the number of quantified proteins is typically lower, around 4,000. Protein identification and quantification also correlate with MS analysis time and sample quality, including factors like lysis conditions and contamination.<\/p>\n\n\n\n<p>For <strong>protein complex analyses<\/strong> (e.g., immunoprecipitations or proximity labeling), only a subset of the proteome is expected\u2014often a few hundred proteins in a typical 90-minute LC-MS\/MS run. We typically use TMT-based quantification for these experiments and to improve the quantification precision and achieve deeper interactome coverage, we usually perform high-pH offline fractionation, analyzing multiple fractions (commonly six).<\/p>\n\n\n\n<p>The total number of identified proteins also depends on the <strong>quality of the FASTA database<\/strong> used. Please note, we cannot identify proteins absent from the database.<\/p>\n\n\n\n<p>In case your protein of interest is not detected, there are several possible reasons. Smaller proteins tend to produce fewer tryptic peptides. The distribution of tryptic cleavage sites along the protein sequence may be uneven. Low expression levels can also limit detection. Additionally, the protein\u2019s cellular localization and the efficiency of lysis can affect whether it is identified. Finally, proteins expressed from other species may not be detected.<\/p>\n\n<\/div>\n<\/details>\n\n<\/div>\n<\/div>\n\n\n<div><!--[vf\/content]-->\n<div class=\"vf-content\">\n\n<div class=\"vf-box vf-box--normal vf-box-theme--primary | vf-u-margin__bottom--400\">\n      <h3 class=\"vf-box__heading\">\n                Ask DocBot!                  <\/h3> \n        <p class=\"vf-box__text\">Try out DocBot to help answer your data analysis questions:<\/p>\n<p class=\"vf-box__text\"><a href=\"https:\/\/wwwdev.ebi.ac.uk\/docbot\/?selected=pcf\" target=\"_blank\" rel=\"noopener\">https:\/\/wwwdev.ebi.ac.uk\/docbot\/?selected=pcf<\/a><\/p>\n<p class=\"vf-box__text\">It\u2019s a new chatbot developed by EMBL-EBI and now trained on PCF documentation to help you get answers to your questions faster. It is still in early development with a focus on data analysis, and we would greatly appreciate your feedback to help us improve it.<\/p>\n<p class=\"vf-box__text\">If you still need our personal support, please don\u2019t hesitate to <a href=\"mailto:pcf@embl.de\">contact us directly<\/a> \u2014 we are always happy to help.<\/p>\n<\/div>\n<\/div>\n<\/div>\n<\/div>\n\n\n\n<p><\/p>\n","protected":false},"excerpt":{"rendered":"","protected":false},"author":15,"featured_media":0,"parent":0,"menu_order":0,"comment_status":"closed","ping_status":"closed","template":"template-title-left-aligned.php","meta":{"_acf_changed":false,"footnotes":""},"embl_taxonomy":[],"class_list":["post-22995","page","type-page","status-publish","hentry"],"acf":[],"embl_taxonomy_terms":[],"_links":{"self":[{"href":"https:\/\/www.embl.org\/groups\/proteomics\/wp-json\/wp\/v2\/pages\/22995","targetHints":{"allow":["GET"]}}],"collection":[{"href":"https:\/\/www.embl.org\/groups\/proteomics\/wp-json\/wp\/v2\/pages"}],"about":[{"href":"https:\/\/www.embl.org\/groups\/proteomics\/wp-json\/wp\/v2\/types\/page"}],"author":[{"embeddable":true,"href":"https:\/\/www.embl.org\/groups\/proteomics\/wp-json\/wp\/v2\/users\/15"}],"replies":[{"embeddable":true,"href":"https:\/\/www.embl.org\/groups\/proteomics\/wp-json\/wp\/v2\/comments?post=22995"}],"version-history":[{"count":132,"href":"https:\/\/www.embl.org\/groups\/proteomics\/wp-json\/wp\/v2\/pages\/22995\/revisions"}],"predecessor-version":[{"id":29189,"href":"https:\/\/www.embl.org\/groups\/proteomics\/wp-json\/wp\/v2\/pages\/22995\/revisions\/29189"}],"wp:attachment":[{"href":"https:\/\/www.embl.org\/groups\/proteomics\/wp-json\/wp\/v2\/media?parent=22995"}],"wp:term":[{"taxonomy":"embl_taxonomy","embeddable":true,"href":"https:\/\/www.embl.org\/groups\/proteomics\/wp-json\/wp\/v2\/embl_taxonomy?post=22995"}],"curies":[{"name":"wp","href":"https:\/\/api.w.org\/{rel}","templated":true}]}}