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Peptide Synthesis Introduction
 

      Solid Phase Peptide Synthesis (SPPS), which was established by the 1964 Nobel Prize in Chemistry winner, Robert Bruce Merrifield, is now the standard method for laboratory synthesis of peptides. The peptide synthesizer at ICOB, Liberty Blue, is an innovative microwave-based platform. The microwave generates an electromagnetic field with high intensity in the center and homogenous energy distribution in the cavity. The polar nature of peptide backbone and reagent mixtures will cause these reaction constituents to align with the alternating electric field from the microwave. This alternating alignment will induce random motion of the peptide chain, break up aggregation and free up the N-terminus of the peptide. The two key steps in peptide synthesis are deprotection and coupling reactions, which are both efficiently assisted by the microwave. Thus, microwave irradiation can facilitate the synthesis of difficult peptides. The protocols for many peptide modifications are well-established in our core Lab, including amidation, acetylation, biotin and fluorescent labels (either at the N- or C-terminus), challenging phosphorylation, cyclization and methylation of Arg or Lys.

微波打開聚合現象
Microwave helps in breaking up aggregation

Applications at Peptide Synthesis Core, ICOB


Example


MW= 1659.83 (17-mer peptide)
Scale: 0.05 mmole
Resin:
 Amide resin
Deprotection: 20% Piperidine w/ 0.1M Oxyma in DMF
Coupling: DIC/Oxyma ; 5-fold excess, Double Coupling for all amino acids
Time: 2.5 hours
Peptide Quantity: 69 mg
Crude Purity: ~95 %

 

 

Microflex LRF, MALDI-TOF CEM  Liberty Blue
EP3

 

Peptide Synthesis Equipment    Manager: June Ching-Chun Lin     Tel-9524
 

Items

Equipment

Brand/Model

Applications

Link

1 Automated Microwave Peptide Synthesizer CEM Liberty Blue™ Automated Microwave Peptide Synthesizer

-Peptide-based immunogens, hormones, etc.
-Enzyme–substrate interactions, ELISA-based analysis/Cell-based binding assay
-Peptide drugs (e.g. ligand targeted liposome for cancer therapy)
-Cell-penetrating peptide (CPP) synthesis

Link

on-line reservation system

2100 Bioanalyzer peptide solubility

2

MALDI-TOF MS

Bruker Microflex LRF MALDI-TOF (Matrix-assisted laser desorption/ ionization time-of –flight)

Molecular weight determination

GeneSpring GX 7.2Link

MJ Real-time PCRon-line reservation system

MJ Real-time PCRPI/MW tool

3

2100 Bioanalyzer

Agilent 2100 Bioanalyzer

The Agilent Bioanalyzer utilizes microfluidics chips for determining RNA, DNA and protein quality, assessing labeled RNA or fragmented RNA size distributions, and genotyping by PCR. We currently only provide RNA QC services.

2100 BioanalyzerUser Manual

2100 BioanalyzerRNA Quality Control Request Form

2100 BioanalyzerDNA Sizing and Analysis Request Form

2100 BioanalyzerProtein analysis Request Form

4

Speed Vac
Thermo Savant
SC210A Speed Vac Plus
The SpeedVac concentrator dries and concentrates samples via centrifugal evaporation.

Speed VacSpeed Vac

 
Peptide Synthesis Rules    Manager: June Ching-Chun Lin     Tel-9524
 
Service object
 

We provide peptide synthesis services to all ICOB researchers and their collaborators.

   
Usage protocol
 
Peptide Synthesizer (Liberty Blue)
 
  1. Please submit your request via the on-line reservation system. Any cancellations should be confirmed by a message to the return email prior to the start of synthesis.
  2. The peptide sequence should be provided in standard notation (N- to C-terminal); any special modifications should be discussed with the manager before submission.
  3. Download and fill out a request form with PI’s signature. The completed form should be delivered to the Peptide Synthesis Core Facility (Rm.401-5).
  4. Rate: NT$350/residue at 0.1 mmol regular scale, HPLC purification fee: NT$2,000; special amino acids, fluorophore labeling, biotinylation, phosphorylation and other modifications are available at extra charge. Please inquire.
  5. Users may only access the services in accordance with the rule
   
MALDI-TOF Microflex LRF
 
  1. Please submit a request via the on-line reservation system. Any cancellations should be confirmed by a message to the return email account.
  2. We provide hands-on training courses. All users MUST book a time at least one week before the course. With the exception of standards and MALDI matrices, users should prepare their own samples and consumables for experiments.
  3. Users should take a certification test within two weeks after training. Independent access to equipment is only allowed if the user has passed the certification; if not, please re-attend the training for qualification.
  4. It is also the responsibility of all users who are working in the laboratory to immediately alert the lab manager to any instrument failure or questions. DO NOT CHANGE ANY SETTINGS OR RECONFIGURE ANY PART OF THE INSTRUMENTS.
  5. ALL activity on the instruments MUST be logged. Be sure that all equipment and work areas are carefully cleaned; EVERYTHING you may have brought into the lab, such as gloves, tubes, tips, samples, papers and bottles should be cleared away.
  6. The MALDI-TOF MS Facility is only open during regular office hours.
  7. Users may only access the services in accordance with the rules.
  8. Rate: NT$100/ hr
 
 

128 Sec.2 ﹐Academia Rd ﹐Nankang ﹐Taipei 115 Taiwan ﹐R.O.C Tel: 02-27899515 Fax: 02-27858059

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