FAQ – Technical

Is there any contamination from the silicone seal?
We tried to find silicone oligomers but have never seen any in the mass spectrum.

How does the sensitivity of the DigestPro MSi compare to the DigestPro MS/DigestPro 96?
The principle of operation as well as most of the parts are identical and the digestion sensitivity is therefore similar. The DigestPro MSi is a bit slower as the DigestPro 96 as it has only one probe, but most of the time is reaction time anyway.

How is contamination avoided?
Gel slices are held in flow-through reaction vials. Reagents are exchanged by nitrogen pressure. The gel slices and peptides resulting from digestion are never touched by a pipetting probe.

How is evaporation controlled in the DigestPro MSi?
The reaction vials are covered by a sealing membrane throughout the procedure. Only a small hole is open for reagent delivery.

How does the DigestPro MSi control the temperature?
The 96 well reaction plate is held in a thermostated rack which can be heated to temperatures of up to 70°C.

Are all samples treated the same?
Standard protocols employ the same procedure for all samples but some variation can be programmed.

What is the time required for a run?
A complete run with washing, reduction, alkylation, rehydration, digestion and peptide elution takes about 10 hours.

Is there positive sample identification?
Samples are introduced in the DigestPro MSi in 96 well plates and digests are obtained in the same format without re-arranging.

Does the DigestPro MSi have to be operated in a fume hood?
No, the instrument comes with a self-contained cabinet. An exhaust tube leading to a ventilation system must be connected to an outlet provided.

Does INTAVIS supply reagents?
As high quality buffers are available from several sources, INTAVIS does not supply reagents. However, we supply reaction and collection plates as well as other consumables.

What determines the limit of detection in the DigestPro MSi?
The detection limit is determined by the signal-to-noise ratio in the mass spectrum. Factors like contamination by ubiquitous proteins or adsorption of peptides to plastic surfaces play a major role. Sample enrichment using Zip Tips as used in the DigestPro MSi has been shown to enhance the sensitivity significantly.

What volumes can be handled by the DigestPro MSi?
The reaction vials have a volume of 200 µl. Wash steps use 60 to 100 µl, the digestion is performed in 15 µl. Volumes down to 10 µl can be dispensed without touching the gel slices. In the MALDI procedure a different needle is used which can handle volumes between 0.5-100 µl.

 FAQ – Application

What is the minimum amount of protein identified with theDigestPro MSi?
As this depends on too many variables we specify the sensitivity as follows:Standard proteins like BSA can be detected after digestion if 0.5 pmol were loaded onto a gel and processed. As a rule of thumb:

  • Proteins visualized with Coomassie Blue can be identified if they are in a database.
  • Spots or bands from silver gels work in many but not all cases.

Does INTAVIS you supply reagents?
As high quality buffers are available from several sources, INTAVIS does not supply reagents. However, we supply reaction and collection plates as well as other consumables.

Can the DigestPro MSi be used for other chemical derivatization?
As long as the principle for reagent incubation and exchange is the same the DigestPro MSi can be programmed for other derivatization reactions.

Can other enzymes than trypsin be used?
The DigestPro MSi has four reservoir positions for enzymes. If the protocols are compatible these can even be used in the same run.

Can the DigestPro MSi be used to digest proteins in solution?
As soluble proteins not contained in a gel matrix can not be retained in the flow-through vials they can not be processed like gel pieces. However, there are protocols for in-solution digestion available using our collection plate to run the whole application.

 FAQ – Software

Does the DigestPro MSi require a computer?
The DigestPro MSi is operated by a PC software that comes with the system. Operation of the instrument and adaptation of protocols is performed by the Windows-based software with an intuitive graphical interface.

Does the DigestPro MSi produce a log of operation?
The complete operation for each pipetting step including time of execution and details about the single steps will be recorded in designated folders on the PC. The log file file containing the information is created during each run.

Can I program my own protocols?
Yes. We recommend to start with the predefined template protocols as most parameters have been set to their optimum in these protocols.

How difficult is progamming of the DigestPro MSi compared to the previous model?
The programming has been simplified. A powerful Windows compatible graphical software used on a PC provides full access to the relevant parameters. New protocols are easily created and stored on a PC.