Peptide mapping against sequence

Amino acid sequence confirmation by peptide mapping

Are you looking to confirm your protein sequence? Alphalyse provides standardized peptide mapping as a fast and convenient service. Your sample is digested with trypsin, cleaving protein sequences after Arg and Lys residues. The resulting peptide mixture is then analyzed by either MALDI-MS/MS or nano LC-MS/MS. The observed peptides are mapped to the expected amino acid sequence.

Choose your service

Our Fast Protein Analysis Services are without customization. You get quality results in easy to understand standard reports, at fixed prices.

20% to 100% coverage

Peptide mapping by MALDI-MS/MS

  • Reduction/alkylation of cysteines, in-gel trypsin digest
  • MALDI-MS/MS peptide analysis
  • Mapping against custom sequence
Per sample
Results in 4-8 days
50% to 100% coverage

Peptide mapping by nano LC-MS/MS

  • Reduction/alkylation of cysteines, in-solution trypsin digest
  • NanoLC-MS/MS peptide analysis
  • Mapping against custom sequence
Per sample
Results in 10-20 days
Service includes
  • We do in-gel trypsin digest and peptide mapping by MALDI-MS/MS or nano LC-MS/MS.
  • We compare with your theoretical amino acid sequence.
  • You obtain a sequence coverage map.
  • You obtain calculated sequence coverage in % (generally 20-100%).
  • Report is delivered within 4-8 days.

Protein sequence coverage map

Our customers typically use this analysis for the following applications:

Map your peptides and confirm them to a theoretical amino acid sequence

Verification of protein sequence of recombinant protein expression

Looking for N- and C-terminal peptides

Analysis of degradation products

Sample requirements

Peptide mapping by mass spectrometry requires a pure protein in sufficient amounts to obtain good data. It is important that samples are prepared in a clean laboratory to avoid contamination with human keratin.

Samples for our PICK ‘n POST peptide mapping can only be delivered as In-gel samples

In-gel samples:
Proteins from 1D SDS PAGE and 2D PAGE gels can be submitted for peptide mapping.
Run your 1D/2D gel

Stain the gel with Coomassie Blue, SYPRO Ruby or Silver stain
Silver stain – It is essential to use a staining protocol or kit that has been optimized for MS analysis. We recommend using one of the following kits:
ProteoSilver Plus, Sigma (Product # PROTSIL1 or PROTSIL2)
Dodeca Silver Stain, BioRad (Product # 161-0481 or 161-0480)
Or use the Alphalyse silver staining protocol

Do not destain silver-stained samples!

Cut out the bands/spots of interest.

Technical details

There are many pitfalls in peptide mapping including incomplete digestion, non-reproducible digestion conditions, non-specific cleavages and non-reproducible chromatography. We at Alphalyse have years of experience in selecting the best digest conditions, ensure enzyme activity and specificity, and performing the mass spec analysis with robust and reproducible protocols. We always include standards that has to fulfill preset criteria to approve your analysis results.

The sequence coverage using a single in-gel digest will typically be between 20 and 100% dependent on the protein amount and the specific protein sequence. Our standard PICK ‘n POST™ analysis is carried out by digesting your in-gel protein sample using Trypsin which cleave the amino acid sequence after Arg and Lys residues . The resulting peptide mixture is analyzed by either MALDI MS/MS or nano LC-MS/MS, and observed peptides are mapped to the expected amino acid sequence. We provide you with a peptide coverage map, illustrating exactly which amino acid sequences that were identified. We also calculate the sequence coverage in percentage. The sequence coverage is typically higher for nanoLC-MS/MS analysis compared to MALDI-MS/MS analysis.

Have questions? We'd love to help.

We're always eager to help. Please send us your question or request and we will get back to you as soon as possible.

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