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Peptide Library Frequently Asked Questions (FAQ)

How are PEPscreen peptides synthesized?

PEPscreen peptides are synthesized on proprietary, state-of-the-art robotic platforms utilizing optimized protocols based on Fmoc-chemistry.


What are the capabilities of the synthesis platforms?

Each platform can simultaneously synthesize hundreds of peptides. Each peptide set can have all unique amino acid sequences and varying peptide lengths (6 to 20 amino acids). Our manufacturing lab has the capacity for synthesizing and processing thousands of peptides per week.


Are modified peptide sequences compatible with the PEPscreen platform?

Yes, we offer a comprehensive portfolio for modifications available at the N-terminal, Internal and C-terminal sequence positions. Available Modifications. Contact peptides@sial.com should you have a special modification request not listed.


What quality control (QC) is performed on the peptide libraries?

ALL peptides are analyzed by MALDI-TOF mass spectrometry (MS) to confirm the correct molecular weight.  The MS profiles are saved as pdf files on a CD and shipped with the peptide library shipment.  Performing and providing the MS analysis ensures the delivery of high quality peptides.


How do you guarantee the quality of PEPscreen peptides?

Sigma has 2 quality criteria that each peptide must reach in order to meet our quality expectations.  

  1. MS criteria: the theoretical molecular weight of the peptide must be found as one of three major ions on the mass spectrometry profile.

  2. Gross weight criteria: the final weight of the peptide must meet the minimum weight guarantee (either 0.5mg or 2mg, depending on the amount ordered)

Should a peptide not meet these criteria, the peptide is synthesized a second time.  If the peptide meets the quality requirements, the product is shipped.  If the remade peptide does not meet the quality requirements, the peptide is marked as ‘failed’.  Failed peptides are shipped to the customer as there is likelihood that the peptide will still provide qualitative value in the full peptide set. However, the customer is not invoiced for any peptides that are categorized as ‘failed’ (not meeting our 2 quality criteria).  Due to the robustness of the chemistry for synthesizing PEPscreen peptides, failed peptides are typically due to the sequence composition (ex. very hydrophobic peptide sequences).


What counterions are present in the peptides?

Typically, the peptides are supplied with trifluoroacetate counterions (TFA). Upon request, the peptides can also be supplied in acetate  or chloride counterions. It is important to note that because there is no purification step, it cannot be guaranteed that the peptide is free of trifluoroacetate counterions. If the peptide set had been supplied in trifluoroacetate, most of the trifuloroacetate counterions can be exchanged to acetate counterions by resuspending the peptides in 200 ml of neat HOAc, followed by re-lyophilization. This procedure is repeated again. Finally, the peptides are re-dissolved in 50% acetic acid in water and then re-lyophilized. The residual trifluoroacetate concentration is usually too low to be a factor in most research applications.


What average purity can I expect for peptides synthesized on the PEPscreen platform?

PEPscreen peptides are supplied unpurified (crude) and the purity of the individual peptides in a set can vary considerably based on the amino acid composition and length of the different peptides in the set. Controlled experiments show that sets of 96 unique sequences containing 10, 15, and 20 residues yield average purities of 80±13%, 65±18%, and 50±20%, respectively. The performance of the synthesis platforms are constantly monitored by synthesizing the ACP control peptide (a 10-mer) to yield a minimum purity of 92±5%.


How can I tell the purity of the peptides in my library?

Analytical HPLC (RP-HPLC) can be performed on individual peptides for an additional fee. The analysis is typically performed using C18 reverse phase column (7.5 cm × 4.6 mm, 3 mm) and the peptide is eluted using elution gradient of water (0.1 % TFA), ACN (0.1 % TFA), 5-75 % over 8 minutes. A summary of the purity profiles will be supplied electronically approximately 3 days after the product has shipped.


How are the peptide sets supplied?

The peptides are supplied as lyophilized film coated at the bottom of 1-ml tubes arranged in a 96-tube plate and individually sealed with silicon rubber lids. Individual tubes are used to allow re-sorting of the peptides in a plate, if necessary, as well as to prevent cross contamination since adjacent tubes are not in contact with each other like in well-plates. The peptides are supplied in a coated film to prevent adhesion to the upper sides and the lid of the tubes during transit.


How do I know that there is peptide in the vial?

The coated peptide film is difficult to see and at best appears as a translucent film on the lower 1/3 of the tube.


What does peptide “gross weight” mean? How does it relate to peptide content and peptide purity?

“Peptide gross weight" refers to the actual weighed weight of the peptide. The gross weight includes the correct peptide, residual water, as well as peptide and non-peptide impurities that may be present and contribute to the weight.

"Peptide content" is more specific and refers to the amount of correct peptide relative to everything else that contributes to the gross weight of the sample, such as residual water, peptide impurities, and non-peptide impurities. "Peptide content" is not obtained from weighing the sample. Instead, it is estimated from the combined data from amino acid analysis and from peptide purity. Peptide content is affected by the amount of water and salts present in the peptide sample.

"Peptide purity" refers to the amount of correct peptide relative to all peptide and non-peptide impurities (except water and salts) that are detected by the assay method. Typically, peptide purity is determined by analytical RP-HPLC and detected by the absorbance at 215 nm. Peptide purity is then estimated from the percentage of the peak area of the correct peptide relative to the combined areas of all peaks present, whether these peaks are derived from peptide or non-peptide impurities. Peptide purity is not affected by the presence of water and salts because these species are generally not detected by the assay method.


Why does the amount of peptides vary between tubes in a peptide set?

The final gross weight of the peptides in a set is a result of different factors, such as the length of the peptide sequence, the types of amino acid residues, the efficiency of each coupling step, the solubility in different solvents used in the production process, and the ability to retain water in lyophilized form. These factors act together, however the contributions of each are impossible to assess.


When should I expect to receive my PEPscreen peptide set?

If the peptide set does not require re-synthesis, the peptides may be shipped in as little as 4 business days. If re-synthesis is required, the order typically ships within 7 business days. Delivery time depends on the size of the peptide set being ordered.

General timelines:

1-2 plates (up to 192 peptides)  = 7 business days

3-4 plates (193 - 384 peptides)  = 12 business days

5-6 plates (385 - 576 peptides) = 15 business days

>6 plates (greater than 576 peptides)  = Inquire


What are the recommended storage conditions for a PEPscreen peptide set?

The lyophilized peptide and peptide solutions should be stored at –20 °C when not in use. For long-term storage (over one year), it is recommended that peptides be stored at –80 °C. Lyophilized powders are generally more stable than peptide solution. In addition, freeze-thaw cycles should be minimized, especially for peptide solutions. For periodic use of the peptides, it is highly recommended to prepare aliquots of peptide solutions and thaw only the aliquots needed to for experiment. More