Frequently Asked Questions (FAQs)
If you are planning to use our service for the first time or if you have specific questions regarding sample preparation you might find the answer here:
Peptide Mass Fingerprint/MALDi-TOF-MS based protein identification
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How much sample is required? |
Any band that is visible by Coomassie staining should contain enough material for identification. |
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How do I send the gel band or spot? |
Please send us your gel band or spot, cut directly from your Coomassie/simply blue-dyed gel, in a 1.5ml Eppendorf container |
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Can you pool several bands from a protein to get more protein? |
Generally the pooling of several gel pieces does not lead to better protein identification. Only an improved focusing of the bands or spots, meaning a higher local concentration of protein in the gel, enables better protein identification. |
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How do I get the results? |
You will get your results in form of a result sheet as pdf via email (Result Sheet (Example)) |
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How well separated must the protein be? Can several proteins be identified from one band/spot? |
A good separation is a prerequisite for successful protein identification via MALDI-MS, especially in the case of 1-D gel electrophoresis. Nevertheless, in some cases it is possible to identify several proteins in one spot/band. Because of the heterogeneity of the sample in some cases it may also happen that no protein can be identified. |
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Will all bands be identified? |
This is a question not easily answered, because the success of the analysis depends on many factors. The most important factor usually is the amount of protein available for analysis (see Q1). Other factors are the resolution of the protein spot/ band, possible contaminations, availability in the protein database, age of the gel, size of the protein and the primary sequence of the protein. For a well focused 2-D gel spot with a medium staining intensity (Coomassie®) and a molecular weight above 15kDa from an already sequenced genome, a probability of success of over 95% can be expected. |
LC-ESI MS/MS based Protein Identification
| How much is required for liquid samples? | For liquid samples 0,5-1µg protein should be available with a concentration of 1mg/ml (purity at least 90%, max. one glycosylation site). No Detergents! |
| How do I get the results? | You will get your results in form of a result sheet as pdf via email (Result Sheet (Example)) |
| Is it possible to detect proportion of two proteins? | Based on signal intensity it is possible to compare the proportions of proteins. But this cannot compare with a quantification, thats only a relative proportion. |
| Which rate of success can be expected? | This is a question not easily answered, because the success of the analysis depends on many factors. The most important factor usually is the amount of protein available for analysis (see Q1). Other factors are the resolution of the protein spot/ band, possible contaminations, availability in the protein database, age of the gel, size of the protein and the primary sequence of the protein. For a well focused 2-D gel spot with a medium staining intensity (Coomassie®) and a molecular weight above 15kDa from an already sequenced genome, a probability of success of over 95% can be expected. |
Quantitative Amino Acid Analysis
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How much sample is required? |
At least 250 µg lyophilised protein. For liquid samples is required amount of 0,5 - 1ml (with concentration of 1 mg/ml). |
| How do I send the protein/antibody? |
Please send us your protein or antibody in solution, in a 1.5ml Eppendorf container: Please provide the buffer composition and the sequence (if possible). |
| How do I get the results? |
You will get your results in form of a result sheet as pdf via email (Result Sheet (Example)) |
N-terminal Sequencing
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How much sample is required? |
The sample should be available amount of at least 20-50 µg protein in 1mg/ml concentration. |
| How do I send the protein/antibody? |
You send us your pure protein or antibody in solution with adequate purity, blotted in to a PVDF membrane, in solution or as lyophilisate in a 1.5ml Eppendorf container. |
| How do I get the results? |
You will get your results in form of a result sheet as pdf via email (Result Sheet (Example)) |
| What are the requirements for N-terminal sequencing samples? |
▪ Care has to be taken that the sequencing protein ist not N-terminal blocked (e.g. through Pyroglutamat, Formyl group or acetyl group) |
Intact Mass determination using ESI-QTOF-MS
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How much sample is required? |
Intact mass samples should be available amount of at least 10 µg; optimum 50-100µg protein in 1mg/ml concentration (purity at least 90%, max. one glycosylation site) |
| How do I send the protein/antibody? |
You send us your protein or antibody in solution, > 90% purity, in a 1.5ml Eppendorf container |
| How do I get the results? |
You will get your results in form of a result sheet as pdf via email (Result Sheet (Example)) |
| What is the fault tolerance of the standard analysis method? | ca. 0.01%.mass divergences at optimum requirements (formulation, amount, concentration) |
| Is it possible, to detect proteins with 40 kDa? | For proteins with 40kDa intact mass analysis can be in progress. |
| Is it possible, to detect proteins with 40 kDa, which distinguish only in one amino acid? | Yes. Such proteins can be detect individual (no guarantee for mixtures) |
