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Quality Control Detail Information

Quality Control Equipment:

HPLC – Gilson HPLC, Agilent UHPLC

Mass Spectral Analysis –Bruker Microflex LT MALDI-TOF, Bruker amaZon SL ion trap

Amino Acid Analyzer – Hitachi L-8900 AAA

 

Reagent Preparation:

HPLC Buffers:   A: HPLC grade water with 0.1% TFA; B: Acetonitrile with 0.08% TFA

Mass Spectral Matrix:      10mgs 4 hydroxy-alpha cyano cinnamic acid in 500uL A, 500uL B

                                                Dissolve in eppendorf, spin down pellet, decant and use supernatant

 

Quality Control Protocol:

  1. HPLC Analysis
    Gradient:  available upon request
    HPLC Injection:  approximately 1mg/mL peptide in HPLC grade water, 100uL (100ug) per injection
    UHPLC Injection: approximately 1mg/mL peptide in HPLC grade water, 1uL (1ug) per injection
    * Other solubility techniques may be required for hydrophobic sequences*
    Percent purity based on peak area

  2. Mass Spectral ESI
    Injection:  approximately 1mg/mL peptide in HPLC grade water, 1uL (1ug) per injection
    * Other solubility techniques may be required for hydrophobic sequences*
    Mass to be within +/- 1 dalton of exact molecular weight.

  3. Mass Spectral MALDI-TOF
    Spot 1uL matrix with 1uL /mL peptide solution
    Let air dry, run sample
    Different voltages, ion charges, and setting shown on Mass Spectral Analysis
    Mass to be within 0.1% of exact molecular weight. (Note: if MW is less than 2000 daltons, MW to be within 2 daltons.)

  4. AAA Analysis (If required)
    Sample must be within 20% for acceptable amino acids
    Sequence ratios must be confirmed by analysis
    Note: Cysteine (C), Methionine (M), and Tryptophan (W) residues are destroyed during analysis and will not appear in Amino Acid Composition.  The peptide hydrolysis process converts Asparagine (N) to Aspartic Acid (D) and Glutamine (Q) to Glutamic Acid (E).  Thus, N and Q residues are calculated theoretically as D and E residues, respectively. 

  5. Dilutions (If required)
    Original stock solution concentration was used to determine dilution factor.  A dilution was then made to requested concentration.  If duplicate/triplicate AAA is requested, an average concentration is used to determine dilution factor.

  

Reassay Interval of  Stored Samples:

Every year or each time a lot is aliquoted from bulk storage.

 

Stability Information:

Lyophilized peptides generally have excellent stabilities, often showing little or no degradation after a few years at -20˚ C. Long term storage (>1year) should be at -80˚ C desiccated, medium term storage (1-12 months) should be at -20˚ C desiccated, short term storage (<1 month) may be at 4˚ C.

 

Molecular formula and mass shifts for common 13C and 15N universally-labeled amino acids.


Three Letter Code

Single Letter Code

Molecular formula of 13C/15N universally-labeled free amino acid

Molecular formula of 13C/15N universally-labeled amino acid residues

Mass shift relative to unlabeled

Ala

A^

(13C)3H7(15N)O2

(13C)3H5(15N)O

(+4)

Arg

R^

(13C)6H14(15N)4O2

(13C)6H12(15N)4O

(+10)

Asn

N^

(13C)4H8(15N)2O3

(13C)4H6(15N)2O2

(+6)

Asp

D^

(13C)4H7(15N)O4

(13C)4H5(15N)O3

(+5)

Cys

C^

(13C)3H7(15N)O2S

(13C)3H5(15N)OS

(+4)

Gln

Q^

(13C)5H10(15N)2O3

(13C)5H8(15N)2O2

(+7)

Glu

E^

(13C)5H9(15N)O4

(13C)5H7(15N)O3

(+6)

Gly

G^

(13C)2H5(15N)O2

(13C)2H3(15N)O

(+3)

Ile

I^

(13C)6H13(15N)O2

(13C)6H11(15N)O

(+7)

Leu

L^

(13C)6H13(15N)O2

(13C)6H11(15N)O

(+7)

Lys

K^

(13C)6H14(15N)2O2

(13C)6H12(15N)2O

(+8)

Met

M^

(13C)5H11(15N)O2S

(13C)5H9(15N)OS

(+6)

Phe

F^

(13C)9H11(15N)O2

(13C)9H9(15N)O

(+10)

Pro

P^

(13C)5H9(15N)O2

(13C)5H7(15N)O

(+6)

Ser

S^

(13C)3H7(15N)O3

(13C)3H5(15N)O2

(+4)

Thr

T^

(13C)4H9(15N)O3

(13C)4H7(15N)O2

(+5)

Tyr

Y^

(13C)9H11(15N)O3

(13C)9H9(15N)O2

(+10)

Val

V^

(13C)5H11(15N)O2

(13C)5H9(15N)O

(+6)

 

Testimonials

"We have no reason to use any other antibody company at this point! We are very pleased with your work. We realize the difficulty in obtaining high quality phosphorylated antibodies so we desire to stick with NEP. Thank you for your work on this!"
Beth A. PhD - Florida State University