We use the Intavis Model MultiPep multiple peptide synthesizer to make up to 192 peptides simultaneously. The synthesis scale is 1-10umol, with 5umol being the most common. This service is ideal for peptide library and screening experiments. Applications include protein binding interactions, epitope mapping, enzyme inhibition and immunodiagnostics. Starting with tentagel resin, the peptide is built "backwards," from the C-terminus to the N-terminus. After synthesis, the peptide is cleaved from the resin, yielding approximately 2.5µmoles of the crude form of the peptide. Generally, this type of synthesis yields crude peptides with 80-85% purity, but HPLC purification can be done if higher purity is desired. Average peptide lengths are 10-20 amino acids. In a typical run, 0-5% of the peptides fail to synthesize properly due to difficulties in coupling caused by the specific amino acid sequence of the peptide. No feedback to adjust chemistry during run but even 50mers are achievable. Runs start on the 1st and 3rd Thursday of each month.
The peptide, in both crude and purified forms, is given out as dry powder . Mass spectrometry data is provided as well (HPLC data is provided upon request).
To determine how much peptide is present:
1. Assume that 50% of the synthesis scale is present for each peptide.
2. Assume that about 75% by weight is peptide, with the remainder being H20 and TFA salt.
3. If there is Tyr, Trp, Phe, or Cys in the sequence, you can use the extinction coefficients for those amino acids to calculate the amount present. (@ 280nm, eY=1280, eW=5690, eC=120, eF=2 e=extinction coefficient in units of Lmol-1cm-1)
epeptide=aeY + beW + ceC + deF, where a=# Y in sequence, b=#W, c=#C, d=#F
Use c=A/(epeptide*l), where c=molar concentration of peptide in mol/L, A=absorbance at 280nm, and l=path length of cuvette in cm
4. Amino acid analysis is the most accurate method. We can help you interpret the AAA data if you like. We suggest these analysis labs:
|AAA Laboratory (Lowell H. Ericsson)
6206 89th Avenue SE
Mercer Island, WA 98040-4512
fax 206 236-6305
AAA Service Labs, Inc
|AAIPharma Services Headquarters
2320 Scientific Park Drive
Wilmington, North Carolina 28405
Tel: (800) 575-4224
|Dr. Mary K. Young
Director Mass Spectrometry and Protein Microsequencing Core Facility
BRI / City of Hope National Medical Center