Owner's manual

[ Care and Use ManUal ]
BioSuite Peptide Analysis Columns 3
2. ThedatainTable1wasgeneratedusingaBioSuiteC
18
, 3 μm PA-A
4.6mmx250mmanda2.1mmx250mmColumn.
 • Proportionatelylesssystembackpressurewillbegeneratedusing
BioSuite C
18
, 3.5 µm PA-B columns that contain 3.5 μm material
compared to the 3 μm material contained in the BioSuite C
18
,
3 μm PA-A column.
 • Proportionatelylesssystembackpressurewillbegeneratedusing
 BioSuitePeptideAnalysisColumnsoflengthslessthan250mm.
3. RecommendedpHRange:2-8.
4. RecommendedOrganicSolvent/AdditiveConcentrationRange:
 • BioSuiteC
18
3μmPA-A:0-100%acetonitrile(CH
3
CN) 
 Upto0.1%formicacid(FA)orupto0.025%trifluoroaceticacid
 (TFA)
 • BioSuiteC
18
3.5μmPA-B:0-100%CH
3
CN
 Upto0.1%TFA
5. RecommendedTemperatureRange:20-50°C
Note: As indicated above, reduce flow rate when operating at ambient tem-
peratures may be required to avoid excessive column pressure on 250 mm
i.d. columns.
6. Recommended Cleaning Solvents/Procedures: A shift in retention or
resolution may indicate contamination of the column. Flushing with
aneatorganicsolvent(e.g.,100%CH
3
CNor80%Isopropanol/20%
CH
3
CN)isusuallysufficienttoremovethecontaminants.
7. Recommended Storage: For overnight storage, lower column tempera-
turetoambient(e.g.,from40°Cto25°C)andcontinuouslyflush
thecolumnwiththemobilephaseat10-20%ofthemaximumrecom-
mendedflowrate.Ifthecolumnisnottobeusedforseveraldays,
lowercolumntemperaturetoambientandstorethecolumnin100%
acetonitrile(CH
3
CN)withoutandeluentadditivessuchasformicacid
orTFA.Donotstorecolumnsinbufferedeluents.Ifyoumobilephase
contained a buffer salt, flush the column with 10 column volumes of
HPLCgradewatersbeforereplacingtheeluentwith100%CH
3
CN for
storage. Failure to perform this immediate step could result in precipi-
tationofthebuffersaltinthecolumnwhen100%CH
3
CN is introduced.
Completely seal the stored column to avoid evaporation and drying out
the bed.
GeneralConsiderationstoMaximizePerformanceofyourBioSuiteC
18
3 μm
PA-A and C
18
3.5 μm PA-B Columns.
II. sample preparatIon
1. Sample impurities often contribute to column contamination.
2. Itispreferabletopreparethepeptidesampleinthemobilephaseora
solventthatisweaker(lessorganicmodifier)thanthemobilephase.
3. Ifthepeptidesampleisnotdissolvedinthemobilephase,ensurethat
the sample, solvent and mobile phases are miscible in order to avoid
sample and/or buffer precipitation.
4. Filtersamplewith0.2μmmembranetoremoveparticulates.Ifthe
sample is dissolved in a solvent that contains an organic modifier
(e.g.,acetonitrile,methanol,etc.)ensurethatthemembranematerial
does not dissolve in the solvent. Contact the membrane manufacturer
withsolventcompatibilityquestions.
III. mobIle phase ConsIderatIons
Prevent air bubbles from entering the column during its installation, use,
and storage since this may cause degradation of column performance through
theformationofchannelsinthepackedbed.Mobilephasesmustbethor-
oughly degassed before use. This can be accomplished by vacuum filtration,
heliumspargingorbyin-linevacuumdegassing.Inadditiontodegassing
the solvent, vacuum filtration of the solvent will also prevent small particles
frompluggingthecolumnfrit.Youcanuse0.20µmor0.45µmmembrane
tofilteraqueousandaqueous/organicmobilephases.
Note: Consult with filtration membrane manufacturer for details on solvent
compatibility.
Note:Use high quality reagents, HPLC grade water, and HPLC grade solvents
for preparing eluents. The useful column lifetime is a function of numerous
factors including: the cleanliness and composition of the mobile phase and
the sample; the flow rate and pressure used; and the temperature. Refer to
the section below about “Cleaning” for information on extending column life.
Note: Cleaning, however, is not effective when the column is damaged by
irreversible sample adsorption, channeling, or exposure of the packing mate-
rial to excessive heat or shock.