BIOCLASS COLUMNS Table of Contents 2 Key Advantages of HALO Fused-Core Columns 3 Selecting the Appropriate Pore Size 4 HALO Columns for Small Molecule Separations 6 HALO BioClass Columns 8 HALO UHPLC and HPLC Guard Columns 9 Column Part Number Listing HALO and Fused-Core are registered trademarks of Advanced Materials Technology, Inc. Discover the Advantages of HALO and HALO BioClass Fused-Core ® Columns 2015 CATALOG
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2015 CATALOG - Labicom...• ~ 20% lower back pressure than most commercially available sub-2 µm non-core columns HALO 2.7 • Comparable resolution and peak capacity to sub-2 µm
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B I O C L A S S C O LU M N S
Table of Contents
2 Key Advantages of HALO Fused-Core Columns
3 Selecting the Appropriate Pore Size
4 HALO Columns for Small Molecule Separations
6 HALO BioClass Columns
8 HALO UHPLC and HPLC Guard Columns
9 Column Part Number Listing
HALO and Fused-Core are registered trademarks of Advanced Materials Technology, Inc.
Discover the Advantages of HALO and HALO BioClass Fused-Core® Columns
2015 CATALOG
KEY ADVANTAGES OF HALO FUSED-CORE COLUMNS
2 |
PERFORMANCE VS. NON-CORE COLUMNSHALO 2• Superior efficiency compared to many popular sub-2 µm
non-core columns
• ~ 20% lower back pressure than most commercially available sub-2 µm non-core columns
HALO 2.7• Comparable resolution and peak capacity to sub-2 µm
non-core columns at half of the back pressure under the same conditions, or…
• Two-fold higher throughput at twice the flow rate vs. sub-2 µm non-core columns at similar pressures
HALO 5• Provides comparable efficiency and resolution to 3 µm
non-core columns for HPLC separations at 50% lower back pressure
HALO BioClass• Comparable to or better than performance of sub-2 µm
non-core columns for bioseparations at 1/3 to 1/2 of the back pressure
• Outperforms legacy non-core columns in terms of peak shape, peak capacity and recovery
HALO FUSED-CORE PERFORMANCEHigh Speed Separations• Smaller reduced plate heights lead to high efficiencies;
narrower and taller peaks, for improved resolution and lower detection limits (LODs and LOQs)
• Flat van Deemter plot and higher linear velocity optimum (Figure C, page 3) allow higher flow rates with minimal column efficiency loss
High Resolution Separations• High efficiency with longer geometries (100, 150, 250 mm)
provides greater resolving power for challenging applications
• Lower back pressure permits columns to be used in series for the most demanding UHPLC and HPLC separations
Excellent Ruggedness and Reproducibility • Less plugging, longer usable column lifetime and greater
uptime due to larger porosity frits (vs. sub-2 µm totally porous (non-core) columns)
- 2 µm frits for HALO 2.7 and HALO 5 - 1 µm frits for HALO 2 vs. 0.2–0.5 µm frits for sub-2
µm non-core columns
• Excellent column-to-column and lot-to-lot reproducibility
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HALO COLUMNS FOR SMALL AND LARGE MOLECULES: SELECTING THE APPROPRIATE PORE SIZEHow to Choose the Right Pore Size?
• Match the column pore size according to the range of molecular weights (MWs) of the analytes in your sample (Table A)
• Small molecules (< 5000 Da) are usually analyzed using HALO 90 Ångstrom columns
- Packing materials with smaller pores have greater surface area, which allows improved retention and loading capacity for lower MW analytes
- When an analyte is too large for the pores, restricted diffusion can occur, which can lead to peak broadening and reduced retention (Figure A)
• For macrocyclic antibiotics and biomolecules such as peptides and proteins, use larger pore sizes such as 160 Å HALO Peptide and 400 Å HALO Protein columns (Page 6)
Peptides (in RT order): Leu-Enk, Bovine Insulin, Human Insulin, Cytochrome C, Lysozyme
Cyt C
Cyt CLyso
Lyso
0 1 2 4 5 7 8 10 11 13 143 9 126 15
Figure A. In this example, broad peaks are observed in the upper chromatogram for bovine and human insulin, cytochrome C and lysozyme (MWs > ~5700 Da) chroma-tographed using a HALO 90 Ångstrom column.This broaden-ing is due to restricted diffusion using the smaller pore column. However, those same four peaks are much narrower and taller in the lower chromato-gram using the 160 Ångstrom HALO Peptide ES-C18 column.
TEST CONDITIONS:
Column: 4.6 x 100 mmPart Number: HALO 2.7 C18: 92814-602 HALO 2.7 Peptide ES-C18: 92124-602Mobile Phase A: 90/10 water/acetonitrile/0.1% TFAMobile Phase B: 30/70 water/acetonitrile/0.1% TFAGradient: 15% B to 50% B in 15 min.Flow Rate: 1.5 mL/min. Temperature: 30 ˚CDetection: UV 220nm, VWDInjection Volume: 5.0 µLLC System: Agilent 1100
Time (min.)
RESTRICTED DIFFUSION FOR HIGHER MW POLYPEPTIDES ON HALO 90Å
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HALO COLUMNS FOR SMALL AND LARGE MOLECULES: SELECTING THE APPROPRIATE PORE SIZE
HALO COLUMNS FOR SMALL MOLECULE SEPARATIONS
Table B. HALO Small Molecule Column Specifications
USP Designation
Particle Size(s) (µm)
Carbon Load (%)
Surface Area (m2/g)
Low pH Limit
High pH Limit
Max. Temp. Lower pH
Limit
Max. Temp. Upper pH
LimitEndcapped
C18 L12
2.75
7.57.75.4
12013590
2 9 60 40 Yes
C8 L72
2.75
4.85.43.7
12013590
2 9 60 40 Yes
Phenyl-Hexyl L112
2.75
6.37.15.2
12013590
2 9 60 40 Yes
PFP L432
2.75
5.35.53.9
12013590
2 9 60 40 Yes
ES-CN L102
2.75
3.53.52.5
12013590
1 8 80 40 Yes
RP-Amide L602
2.75
7.38.25.5
12013590
2 9 60 40 Yes
HILIC L32
2.75
N.A.12013590
1 8 60 40 N.A.
Penta-HILIC Pending2
2.75
2.83.22.1
12013590
2 9 60 40 No
Phase Structure Phase Structure
C18 ES-CN
C8 RP-Amide
Phenyl-Hexyl HILIC
PFP Penta-HILIC
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HALO COLUMNS FOR SMALL MOLECULE SEPARATIONS
Table C. HALO Phases: Features and Benefits, Target Analytes and Best Applications
Bonded Phase Features and Benefits Target Analytes Best Applications
C18 C18(octadecyldimethylsilane)
Excellent performance for broad range of analyte polarities
Diverse analytes ranging from polar to non-polar
Uncharged acids and bases, uncharged ion pairs
Analytes differing by an ali-phatic or aromatic group
C8 C8(octyldimethylsilane)
Excellent performance for broad range of analyte polarities
Diverse analytes ranging from polar to non-polar
Uncharged acids and bases, uncharged ion pairs
Analytes differing by an aliphatic or aromatic group
More retention for polar analytes and much less reten-tion for non-polar analytes
Polar and very polar bases, acids and neutrals
Very hydrophobic analytes retained too strongly on C18 phase
Aromatic molecules with electron-withdrawing groups (NO2, COOH, COOR, halogens), heterocycles, benzodiazepines, highly aqueous conditions
RP-Amide C16 Amide
Complementary selectivity to alkyl phases
Enhanced stability for mini-mum bleed and long life
Alcohols, Acids, Phenols, Catechins
Acidic and basic analytes, heterocycles, proton donors and acceptors, highly aque-ous conditions
HILIC None (Bare Silica)
Can be used in HILIC and normal-phase modes
Polar and very polar bases, acids and neutrals, especially with log P<0.5
Enhanced sensitivity and peak shape for LC-MS, analyses of basic analytes in HILIC mode, normal-phase analysis with non-aqueous mobile phases
Penta-HILIC proprietary penta-hydroxy ligand
Ideal for separation of highly polar compounds that are poorly retained in RPLC
Polar analytes with log P values near or less than 0
Polar acids, bases and zwitterions that are not retained or are poorly retained with RPLC
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HALO Protein C4• Stability up to 90˚C
• Can elute very large proteins with good peak shape and recovery
• Compatible with UHPLC and HPLC
• Low LC-MS bleed
HALO Protein ES-C18• Extremely stable up to 90˚C
• Can elute very large proteins with good peak shape and recovery
• Compatible with UHPLC and HPLC
• Very low LC-MS bleed
HALO Peptide ES-C18• Fast, high resolution separations
• Extremely stable up to 90˚C
• High peak capacity
• Rugged, reliable performance
• Use with either UHPLC or HPLC HALO Peptide ES-CN • Same benefits as Peptide ES-C18
• Alternative selectivity to Peptide ES-C18 for peptide mapping and proteomic applications
HALO Glycan • Improved retention of acidic and
zwitterionic analytes
• Very low sensitivity to buffer concentration
• Able to separate isobaric oligosaccharides with different linkages
HALO PROTEIN, PEPTIDE AND GLYCANToday, researchers are keenly interested in both fast and high resolution separations of numerous biomolecules to support the development of novel therapeutic proteins and peptides in pharmaceutical drug development, to advance understanding in modern university laboratories, to characterize protein post-translation modifications, and to fully assess subtle differences in biosimilars and other products of bioengineering and manufacture. HALO BioClass columns have been developed to make such tasks easier and more effective, and are aimed at the following areas of bioseparations:
• Intact proteins, monoclonal antibodies (mAbs), biosimilars, and other large biomolecules such as pegylated proteins, antibody drug conjugates (ADCs), etc.
• Peptide mapping (analysis of enzyme digests) for characterization and monitoring of synthetic protein drugs
• Analysis of therapeutic peptides and peptide biomarkers (protein surrogates) • High resolution separations of complex mixtures of glycans released from N- and O-linked glycoproteins
Currently, the HALO BioClass column family is comprised of the following products. Their features and benefits are described below, and their specifications are shown in Table D.
• Collect strongly retained compounds from the sample and minimizes column fouling
• Ultra-low dispersion, easy to use, operate at pressures up to 1000 bar
• Finger-tight, direct-connect units that auto-adjust to any column with a 10–32 inlet port
• Easily replace guard cartridge without removing guard holder from the flow path
• Available for all HALO analytical geometries (2.1, 3.0 and 4.6 mm ID) and phases
See figure below for an exploded view of the HALO guard cartridge and guard holder. Please see pages 9–13 for ordering information.
Finger-tight guard cartridge holder, Inlet
Provides easy replacement of guard car-
tridge
Titanium hybrid ferrule
Adds durability for multiple connections
Finger-tight guard cartridge holder, OutletAllows for replacement
of guard cartridge without re-moving the guard holder
from the flow path
HALO guard cartridgeAll HALO phases available
Auto-adjusting zero dead volume (ZDV) end fitting
Ensures optimum, ZDV connection to the analytical column
HALO UHPLC AND HPLC GUARD COLUMNS
The Optimize Technologies EXP Titanium Hybrid Ferrule is Patent Pending. The EXP Holder is patented under PatentNo. 8,696,902 B2. The OPTI- prefix is a registered trademark of Optimize Technologies, Inc.
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The part numbers for HALO 2 columns are presented below. HALO 2 columns are available in the following analytical internal diameters (2.1 and 3.0 mm). Guard columns are also available for HALO 2 columns for these IDs for UHPLC to provide additional protection when necessary.
HALO 2 COLUMNS
DimensionsID x Length (in mm) C18 C8 Phenyl Hexyl RP-Amide PFP ES-CN Penta-HILIC HILIC
ID x Length (in mm) C18 C8 Phenyl Hexyl RP-Amide PFP ES-CN Penta-HILIC HILIC2.1 x 5 91812-102 91812-108 91812-106 91812-107 91812-109 91812-104 91812-105 91812-1013.0 x 5 91813-102 91813-108 91813-106 91813-107 91813-109 91813-104 91813-105 91813-101
94900-001
2 µm, 90 Å
Guard Column Holder
Visit our web site (www.advanced-materials-tech.com) or contact one of our distributors.
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HALO 2.7 COLUMNSHALO 2.7 columns are available in nano, capillary and analytical diameters, as well as in a 10 mm semi-preparative diameter. Guard columns are available for analytical diameters of 2.1, 3.0, and 4.6 mm.
Visit our web site (www.advanced-materials-tech.com) or contact one of our distributors.
HALO 5 COLUMNSHALO 5 columns are available in nano, capillary and analytical diameters, and in a 10 mm semi-preparative diameter. HALO 5 Guard columns are available for analytical diameters of 2.1, 3.0 and 4.6 mm.
Visit our web site (www.advanced-materials-tech.com) or contact one of our distributors.
HALO PEPTIDE COLUMNSThe part numbers are provided below for the nano, capillary, analytical and semi-preparative HALO 2.7 and HALO 5 Peptide columns in both ES-C18 and ES-CN phases. Guard columns are available for 2.1, 3.0 and 4.6 mm internal diameters for UHPLC and HPLC applications, if additional protection is desired.
DimensionsID x Length (in mm) HALO 2.7 Peptide ES-C18 HALO 2.7 Peptide ES-CN HALO-5 Peptide ES-C18 HALO-5 Peptide ES-CN
0.075 x 50 91229-402 91229-404 91529-402 91529-4040.075 x 100 91229-602 91229-604 91529-602 91529-6040.075 x 150 91229-702 91229-704 91529-702 91529-704
0.1 x 50 91228-402 91228-404 91528-402 91528-4040.1 x 100 91228-602 91228-604 91528-602 91528-6040.1 x 150 91228-702 91228-704 91528-702 91528-704
0.2 x 50 91227-402 91227-404 91527-402 91527-4040.2 x 100 91227-602 91227-604 91527-602 91527-6040.2 x 150 91227-702 91227-704 91527-702 91527-704
0.3 x 50 91226-402 91226-404 91526-402 91526-4040.3 x 100 91226-602 91226-604 91526-602 91526-6040.3 x 150 91226-702 91226-704 91526-702 91526-704
0.5 x 50 91225-402 91225-404 91525-402 91525-4040.5 x 100 91225-602 91225-604 91525-602 91525-6040.5 x 150 91225-702 91225-704 91525-702 91525-704
1.0 x 30 92121-302 92121-304 95121-302 95121-3041.0 x 50 92121-402 92121-404 95121-402 95121-4041.0 x 75 92121-502 92121-504 95121-502 95121-504
1.0 x 100 92121-602 92121-604 95121-602 95121-6041.0 x 150 92121-702 92121-704 95121-702 95121-704
2.1 x 20 92122-202 92122-204 95122-202 95122-2042.1 x 30 92122-302 92122-304 95122-302 95122-3042.1 x 50 92122-402 92122-404 95122-402 95122-4042.1 x 75 92122-502 92122-504 95122-502 95122-504
2.1 x 100 92122-602 92122-604 95122-602 95122-6042.1 x 150 92122-702 92122-704 95122-702 95122-7042.1 x 250 92122-902 92122-904 95122-902 95122-904
3.0 x 20 92123-202 92123-204 95123-202 95123-2043.0 x 30 92123-302 92123-304 95123-302 95123-3043.0 x 50 92123-402 92123-404 95123-402 95123-4043.0 x 75 92123-502 92123-504 95123-502 95123-504
3.0 x 100 92123-602 92123-604 95123-602 95123-6043.0 x 150 92123-702 92123-704 95123-702 95123-7043.0 x 250 92123-902 92123-904 95123-902 95123-904
4.6 x 20 92124-202 92124-204 95124-202 95124-2044.6 x 30 92124-302 92124-304 95124-302 95124-3044.6 x 50 92124-402 92124-404 95124-402 95124-4044.6 x 75 92124-502 92124-504 95124-502 95124-504
4.6 x 100 92124-602 92124-604 95124-602 95124-6044.6 x 150 92124-702 92124-704 95124-702 95124-7044.6 x 250 92124-902 92124-904 95124-902 95124-90410.0 x 50 92120-402 92120-404 95120-402 95120-40410.0 x 75 92120-502 92120-504 95120-502 95120-504
10.0 x 100 92120-602 92120-604 95120-602 95120-60410.0 x 150 92120-702 92120-704 95120-702 95120-70410.0 x 250 95120-902 95120-904
Guard Columns, 3-pack
Guard Columns
DimensionsID x Length (in mm) HALO 2.7 Peptide ES-C18 HALO 2.7 Peptide ES-CN HALO-5 Peptide ES-C18 HALO-5 Peptide ES-CN
2.1 x 5 92122-102 92122-104 95122-102 95122-1043.0 x 5 92123-102 92123-104 95123-102 95123-1044.6 x 5 92124-102 92124-104 95124-102 95124-104
94900-001
Visit our web site (www.advanced-materials-tech.com) or contact one of our distributors.
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HALO PROTEIN COLUMNSPart numbers for nano, capillary, analytical and semi-preparative HALO Protein columns in both C4 and ES-C18 phases are provided below. Guard columns are available for HALO Protein columns in 2.1, 3.0 and 4.6 mm IDs for UHPLC and HPLC applications to provide additional column protection when desired.
Dimensions(in mm) HALO Protein C4 HALO Protein ES-C18
0.075 x 50 94319-414 94319-4020.075 x 100 94319-614 94319-6020.075 x 150 94319-714 94319-702
0.1 x 50 94318-414 94318-4020.1 x 100 94318-614 94318-6020.1 x 150 94318-714 94318-702
0.2 x 50 94317-414 94317-4020.2 x 100 94317-614 94317-6020.2 x 150 94317-714 94317-702
0.3 x 50 94316-414 94316-4020.3 x 100 94316-614 94316-6020.3 x 150 94316-714 94316-702
0.5 x 50 94315-414 94315-4020.5 x 100 94315-614 94315-6020.5 x 150 94315-714 94315-702
1.0 x 30 93411-314 93411-3021.0 x 50 93411-414 93411-4021.0 x 75 93411-514 93411-502
1.0 x 100 93411-614 93411-6021.0 x 150 93411-714 93411-702
2.1 x 20 93412-214 93412-2022.1 x 30 93412-314 93412-3022.1 x 50 93412-414 93412-4022.1 x 75 93412-514 93412-502
2.1 x 100 93412-614 93412-6022.1 x 150 93412-714 93412-702
3.0 x 20 93413-214 93413-2023.0 x 30 93413-314 93413-3023.0 x 50 93413-414 93413-4023.0 x 75 93413-514 93413-502
3.0 x 100 93413-614 93413-6023.0 x 150 93413-714 93413-702
4.6 x 20 93414-214 93414-2024.6 x 30 93414-314 93414-3024.6 x 50 93414-414 93414-4024.6 x 75 93414-514 93414-502
4.6 x 100 93414-614 93414-6024.6 x 150 93414-714 93414-70210.0 x 50 93410-414 93410-40210.0 x 75 93410-514 93410-502
10.0 x 100 93410-614 93410-60210.0 x 150 93410-714 93410-702
Guard Columns, 3-packDimensions
(in mm) HALO Protein C4 HALO Protein ES-C182.1 x 5 93412-114 93412-1023.0 x 5 93413-114 93413-1024.6 x 5 93414-114 93414-102
Guard Column Holder 94900-001
HALO GLYCAN COLUMNSHALO Glycan columns are available in 2.1 and 4.6 mm diameters in the following lengths in 2.7 µm particle sizes. Guard columns are available for UHPLC and HPLC applications if additional protection is desired.
Dimensions(in mm) HALO Glycan
2.1 x 50 92922-4052.1 x 100 92922-6052.1 x 150 92922-705
4.6 x 50 92924-4054.6 x 100 92924-6054.6 x 150 92924-705
Guard Columns, 3-packDimensions
(in mm) HALO Glycan2.1 x 5 92922-1054.6 x 5 92924-105