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[ Care and Use ManUal ]
The AccQ•Tag™ Ultra Derivatization Kit is a component of the UPLC®
Amino Acid Analysis Application Solution. This application solution is
an integrated combination of instrumentation, derivatization chemistry,
separation column and eluents, methods, and software. Analysts are
assured of accurate and precise amino acid analyses with the complete
application solution. The use of the AccQ•Tag Ultra Derivatization Kit
without the rest of the application solution is not supported as an amino
acid analysis method because there are significant risks of inaccurate
qualitative and quantitative analysis.
The AccQ•Tag Ultra Derivatization Kit differs from the AccQ•Tag
Derivatization Kit (P/N WAT052880). Although the specified chemical
composition is the same for both kits, the quality control tests for the
AccQ•Tag Ultra Derivatization Kit are executed with UV detection. In
contrast, the quality control tests for the AccQ•Tag Derivatization Kit
are executed with fluorescence detection. Substituting one kit for the
other may result in unexpected interferences that may compromise
sensitivity or introduce ambiguity into peak identification.
AccQ•TAg UlT rA Ov erv iew
The Waters UPLC Amino Acid Analysis Application Solution is pre-column
derivatization and analysis technique. To perform this analysis:
1. Use Waters AccQ•Tag Ultra Reagent Powder (US Patent 5,296,599
and European Patent EP 0 533 200 B1) to derivatize the amino acids.
2. Separate the derivatives using the properly configured Waters
ACQUITY UPLC® System.
3. The reversed-phase separation occurs on the Waters AccQ•Tag
Ultra, 2.1 x 100 mm, 1.7 µm column. The derivatives are eluted
with the specified dilution of Waters AccQ•Tag Ultra Eluent A and
Waters AccQ•Tag Ultra Eluent B.
4. Quantitate the derivatives with UV detection.
5. Process the data using Empower™ software.
For complete details of the instrument configuration and of the separation
and analysis methods, consult the UPLC Amino Acid Analysis Application
Solution System Guide.
cOnT enTs Of K iTThe Waters AccQ•Tag Ultra Derivatization Kit contains five sets of the
derivatizing reagents used for the UPLC Amino Acid Analysis Application
Solution. Each set of reagents includes one vial each of:
• AccQ•Tag Ultra Borate Buffer - Vial 1. The buffer is added to the
samples to ensure the optimum pH for derivatization.
• AccQ•Tag Ultra Reagent Powder – Vial 2A. The reagent powder
is the 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC)
derivatizing reagent. It is shipped dry for maximum stability.
• AccQ•Tag Ultra Reagent Diluent – Vial 2B. This diluent, acetonitrile,
is used reconstitute the reagent for derivatization.
AccQ•TAg UlT rA DerivAT izAT iOn KiT
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[ Care and Use ManUal ]
2
sAmple p repArAT iOn
I. Reconstitute AccQ•Tag Ultra Reagent Powdera. Preheat a heating block to 55 ˚C.
b. Tap vial 2A (AccQ•Tag Ultra Reagent powder) to ensure all reagent
is at bottom of vial.
c. Draw 1 mL of AccQ•Tag Ultra Reagent Diluent from vial 2B.
d. Transfer to vial 2A.
e. Vortex for 10 seconds.
f. Heat on top of heating block, vortex occasionally, until the powder
dissolves. Do not heat longer than 15 minutes.
II. Prepare 100 pmol/μL calibration standard for protein and peptide hydrolysates a. Transfer the Amino Acid Hydrolysate Standard (P/N WAT088122)
from the supplied ampule to a vial appropriate for storage in a freezer.
b. Dilute Amino Acid Hydrolysate Standard to appropriate concentration
by taking 40 µL of standard and adding it to 960 µL of water.
III. Derivatize standards and samplesa. Use a micropipettor to deliver 70 µL of AccQ•Tag Ultra Borate Buf-
fer (vial 1) to a Total Recovery vial (P/N 186000384C).
b. Use a micropipettor to add 10 µL of the calibration standard or dissolved
sample amino acids to the vial. Vortex for several seconds.
c. Use a micropipettor to add 20 µL of reconstituted AccQ•Tag Reagent
to the vial.
d. Cap vial and vortex for several seconds.
e. Let stand for one minute at room temperature.
f. Heat in heating block for 10 minutes at 55 ˚C.
g. Remove from heating block with forceps and place in instrument for
analysis. The resulting standard has a concentration of 10 pmol/µL.
sTOrAge AnD sTAbil iT y• The Waters AccQ•Tag Ultra Derivatization Kit may be stored at
room temperature for up to one year.
• Reconstituted AccQ•Tag Ultra Reagent can be stored at room
temperature in a dessicator with dry dessicant for up to one week.
Note: AccQ•Fluor™ Ultra Reagent reacts with atmospheric water. Seal the
vial tightly. Do not refrigerate because of the risk of condensation.
• Derivatized amino acid samples may be stored at room temperature
for up to one week.
OrDering infOrmAT iOn
AccQ•Tag Ultra Derivatization Kit, 250 analyses 186003836
AccQ•Tag Ultra Column, 2.1 x 100 mm, 1.7 µm 186003837
AccQ•Tag Ultra Eluent A Concentrate, 1 L 186003838
AccQ•Tag Ultra Eluent B, 1 L 186003839
Amino Acid Hydrolysate Standard, 10 x 1mL WAT088122
Sample Tubes, 6 x 50 mm, 72/pkg WAT007571
Total Recovery Vials, 100/pkg 186000384C
Representative Chromatogram: Protein and Peptide Hydrolysates (10 pmol)
AMQ
NH3 His
Arg
Gly Asp Gl
u Thr
Ala
Pro
Ser
Derivitization Peak Cys
Lys
Tyr
Met Val
NVa
ILe
Leu
Phe
M inutes1.00 1.50 2.00 2.50 3.00 3.50 4.00 4.50 5.00 5.50 6.00 6.50 7.00 7.50 8.00
Minutes
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[ Care and Use ManUal ]
Waters Corporation 34 Maple Street Milford, MA 01757 U.S.A. T: 1 508 478 2000 F: 1 508 872 1990 www.waters.com
Waters, AccQ•Tag, AccQ•Fluor, Empower, ACQUITY UPLC, UPLC and The Science of What’s Possible are trademarks of Waters Corporation.
©2012 Waters Corporation. 715001331 Rev D 10/12 VW-PDF
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