[accordion] [accordion_item title=”Can I collaborate with you? I have a great idea for a novel proteomics research project!” active=“no”]Sure! Please contact Jeroen Demmers and arrange a consultation. Also see our website for information on collaborative research projects and services.[/accordion_item] [accordion_item title=”What fees are involved for research projects?” active=“no”]Cost structures need to be negotiated before the work begins. Also take a look at our fees. Please note that all services and projects undertaken will be on a collaborative basis and it is expected that our contributions to your research project will merit co-authorship on subsequent publications.

Charges include costs for reagents only – the Center realizes no profit at all. In contrast, the real costs of analysis are several times that of what we charge you. Data analysis and consultation regarding study design, data interpretation, and publication / public access are provided at no cost.[/accordion_item][accordion_item title=”What is the turnaround time for receiving results for mass spectrometry analysis?” active=“no”]Difficult to say. Some samples need more attention than others. Also, there is a sample queue most of the time. In general, we will let you know when the analysis is finished and send you the results by email, so: don´t call us, we’ll call you.[/accordion_item] [accordion_item title=”I want the best possible results – how do I deliver my samples to your Center?” active=“no”]We try more and more to avoid separation of proteins by SDS-PAGE. Instead, we prefer to receive the samples at the earliest stage in your sample prep procedure as possible. If you have done a co-IP experiment, leave the proteins bound to the beads and bring us the beads; if you want a full proteome coverage of your cells, just bring us the cell pellet after the final extensive washes in PBS or so; if you have tissue samples, bring us the frozen tissue. If you are not sure, talk to us first and we will discuss what’s best for your samples![/accordion_item] [accordion_item title=”Will your staff cut out a protein band from my SDS-PAGE gel?” active=“no”]Yes, we prefer that you bring the Coomassie stained gel and let us cut out your lane or bands. Our experienced staff knows all the tips and tricks to avoid (keratin) contamination of your samples. Also, it is important for us to know what the gel looks like to be able to estimate the total amount of protein.[/accordion_item][accordion_item title=”What protein staining method do you recommend for SDS-PAGE gels used for mass spectrometry results?” active=“no”]Coomassie Blue stained gels are best. Silver staining works well, but the mass spec sensitivity will be somewhat lower.[/accordion_item] [accordion_item title=”Can I cut bands out of my own gel and give you a gel slice?” active=“no”]Yes, you can.. However, we prefer that you drop off the intact gel and mark the bands on the gel scan that need to be cut.[/accordion_item] [accordion_item title=”I have a silver stained gel. Can I still use this for mass spectrometry analysis?” active=“no”]Yes, you can, but be aware that we would identify more proteins from a Coomassie stained gel, even though there appear to be more bands on a silver stained gel.[/accordion_item] [accordion_item title=”What type of mass spectrometers does your lab use?” active=“no”]We use an Orbitrap Lumos, an Orbitrap Fusion, a Q Exactive PLus and two LTQ-Orbitrap XL mass spectrometers (one of them with ETD technology), all of them connected to nanoflow LC systems (Agilent, Eksigent and Waters nanoAcquity UPLC). For student projects we use an old, but still fully functional LTQ linear ion trap. See the Equipment section for details. [/accordion_item][accordion_item title=”I need some basic help with understanding mass spectrometry experiments and interpreting the MS data report of my samples. Who can I talk to?” active=“no”]

All questions are welcome, particularly at the planning stages, before samples are prepared and analyzed. Please contact Jeroen Demmers.

For general questions about:
– Sample handing: see Karel or Dick
– Which buffers are compatible with proteolytic digestion and / or MS analysis: see Karel or Dick
– Protein identification via LC-MS: see Jeroen, Karel or Dick
– Invoices / payments: see Dick

See Jeroen for planning of your experiments and for:
– Your first time with mass spectrometry
– Post-translational modifications
– Complex protein samples
– Contract / collaborative research
– For interpretation of data
– For questions about data format

For follow-up questions about significance of the data analysis For questions about bioinformatics or requests for specific data formats: see Jeroen or Erikjan[/accordion_item] [accordion_item title=”What kind of post-translational modifications can you detect?” active=“no”]Any posttranslational modification can be detected: phosphorylation, methylation, acetylation, ubiquitination, etc. Contact us for details. [/accordion_item] [accordion_item title=”What are the concerns about mapping PTMs in my protein?” active=“no”]You will have two major concerns: 1) Coverage: can the mass spectrometry technique deliver high sequence coverage? With higher coverage, there is a greater statistical chance to detect and identify the peptide containing a modification group. 2) % occupancy of the modification site (stoichiometry): what percent of the protein preparation is modified? If it is low, then the chance of detecting the modified peptide decreases. If the occupancy is high, then this works in favor of identifying a modification site.[/accordion_item] [accordion_item title=”How can coverage be increased for mapping phosphorylation, methylation, ubiquitination, etc. sites? (I understand that high coverage is essential for increasing the chance of mapping a PTM site)” active=“no”]First, note that, due to its unique sequences, every protein has a different potential to be mapped at high sequence coverage. Coverage for all proteins increases by: 1) Starting with a higher amount of protein for MS analysis 2) Making sure that the protein is pure 3) Using multiple MS instruments/techniques for analysis will increase coverage 4) Enriching phosphopeptides using TiO2[/accordion_item] [accordion_item title=”Will glycosylation sites inhibit the identification of my protein?” active=“no”]If purity and concentration of the protein is high, then in most cases MS-based protein identification is still favorable. Attached carbohydrate (CHO) chains, however, can impair MS-based identification of specific peptides through the following mechanism: 1) CHO chains can block trypsin cleavage 2) The variable mass of attached CHO chains can reduce glycopeptide detection. Options: Cleave N-linked CHO chains with N-glycanase (PNGaseF) prior to gel loading and MS analysis[/accordion_item] [accordion_item title=”How much protein do I need in my sample?” active=“no”]Hard to say in general terms. It depends on the specific questions you have: do you need to just identify the proteins or do you need information about PTMs? Is your protein pure or will the peptides of your protein be masked by peptides from background proteins in a complex matrix? What we can say is that pure peptide standards can be identified at sub-femtomole levels using nanoflow LC-MS/MS.[/accordion_item] [accordion_item title=”My protein preparation is not very pure. Can I use it for mass spectrometry analysis?” active=“no”]Yes, you can still use mass spectrometry to identify your protein.[/accordion_item] [/accordion]