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| Updates |
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| Peptide Synthesis FAQs |
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- 1. What is the typical turnaround time for peptide synthesis at Crnbio?
Our typical turnaround time is about three weeks. The turnaround time may vary depending on the peptide length and complexity of synthesis.
- 2. How do you ship peptides? What data will be provided?
All peptides will be lyophilized and shipped in small microcentrifuge tubes (2 ml). Large amounts of peptide may be aliquoted into several tubes. For every single peptide, data sheets containing information such as amino acid sequence, modifications, purity, mass spectrum data, and HPLC data will be provided.
- 3. What is peptide purity?
The purity of Crnbio’s catalog peptides is usually about 95%. This means that 95% of the NET PEPTIDE content (but not total peptide content, see question 11) of the dry powder shipped to you is composed of your target peptide. The other 5% of the PEPTIDE material in your sample is usually composed of the so-called deletion sequences that sometimes co-purify with the target peptide. Deletion sequences are generated during peptide synthesis, when due to the slight inefficiencies of the coupling reaction some amino acids are skipped in some of the synthesized molecules. Purity is usually determined by reverse-phase HPLC.
- 4. How pure does my peptide need to be?
That depends on your specific application. Crnbio can synthesize peptides to 98% purity. Here is a general guideline for peptide purity requirements:
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Purity |
Application |
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>85% |
Immunological applications and polyclonal antibody production |
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>90% |
SAR studies, bioassays |
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>95% |
In vitro bioassays such as ELISA, enzymology, biological activity |
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>98% |
NMR, crystallography, sensitive bioassays |
- 5. What methods do you use to purify the peptide?
HPLC is used for purification.
- 6. What is net peptide content?
It is important to understand the difference between net peptide content and total peptide content. The dry peptide powder shipped to you usually contains not only peptide, but also some other substances such as water, absorbed solvents, counter ions and salts. The total peptide content refers to the weight of this mixture. Net peptide weight indicates the actual weight of the peptide component of your sample. Net peptide content is usually 50-80% of the total peptide weight (also called gross peptide weight) and is usually determined by amino acid analysis or UV spectrophotometry. Net peptide content should not be confused with purity. Purity defines the percentage of the target peptide sequence in the peptide component of your sample.
- 7. How should I store the peptides?
Lyophilized peptides can be stored long-term at -20°C.
- 8. What is the best way to dissolve the peptide?
The solubility of given peptides varies depending on their amino acid sequences and modifications. Crnbio purifies peptides by HPLC using a water and acetonitrile gradient. Here are some general tips for dissolving peptides:
- o Sonication will increase solubility.
o 10% acetic acid in the solvent will help dissolve basic peptides.
o 10% ammonium bicarbonate will help dissolve acidic peptides.
o For peptides with extremely low solubility in aqueous solutions, organic solvents (such as DMSO, isopropanol, methanol, and acetonitrile) should be used first. Once the peptides are completely dissolved, water may be gradually added until the desired concentration is obtained.
- 9. What is a counterion?
Most peptides except those that do not have basic amino acids such as Arg, Lys, His or those with blocked N-termini exist in the form of their salts. Synthetic peptides that are purified by HPLC are usually obtained as TFA salts. Their basic amino acid residues and N-termini are protonated and have trifluoroacetate (CF3COO -) counterions.
- 10. What if there is a problem with the synthesis of my peptide?
Each peptide has its specific characteristics and not all outcomes can be anticipated. If something goes wrong during the synthesis and we cannot deliver your peptide on time, we will inform you as soon as possible.
- 11. What quality control data is provided by Crnbio?
Quality assurance documentation provided with every Crnbio peptide includes mass spectral and HPLC analyses determining composition and purity. Amino acid analysis is available upon request. We also provide storage and handling guidelines.
- 12. In what direction are the peptides synthesized?
Peptides are synthesized from the C-terminus to the N-terminus of the sequence.
- 13. What are the applications of peptide libraries?
Peptide libraries are efficient tools for GPCR ligand screening, protein-protein interaction studies, functional proteomics, nucleic acid binding, enzyme substrate and inhibitor screening, antigen and epitope screening, peptide/protein cross-talk studies, the discovery of signal molecules, and other processes significant to modern drug discovery.
- 14. Are there any requirements for phosphopeptide design?
We recommend that you position the phosphorylated residue no more than 10 residues away from the N-terminus because the coupling efficiency of residues following a phosphorylated residue is significantly reduced.
- 15. Are there any requirements for introducing dye modifications?
We recommend that you add a spacer between the peptide and the dye molecule. This will reduce the chance of the dye affecting peptide folding and binding to receptors. However, if the purpose of the dye labeling is to quantify fluorescence transfer between different structures spacers should not be introduced.
- 16. What is the advantage of capping the N and C termini of the peptide?
Capping will make peptide appear more like native protein. The N terminus can be capped with an acetyl group and C terminus with an amide group. |
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