HALO® BioClass Fused-Core®

Today, researchers are keenly interested in both fast and high-resolution separations of numerous biomolecules. The HALO Fused-Core technology supports the development of novel therapeutic proteins and peptides in pharmaceutical drug development to advance understanding in modern university laboratories, enabling researchers to characterize protein post-translational modifications and fully assess subtle differences in biosimilars and other products of bioengineering and manufacture. HALO BioClass columns have been specifically designed to accomplish these bioseparation goals with a simplified and more effective solution.

With both tailored particle and pore size options, HALO BioClass offers application specific solutions for:

• 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

HALO UHPLC columns are not your typical UHPLC columns. The particles packed into HALO columns are made using groundbreaking Fused-Core® particle technology that produces superficially porous particles (also known as core-shell particles) that make possible the manufacture of very high efficiency columns without the very high pressure of other UHPLC columns. HALO®BioClass columns, thanks to Fused-Core technology, minimize the compromises that must be made between efficiency, speed of separation and column back pressure. It is like combining the benefits of low pressure and high speed that one gets from monolith columns with the high efficiency of UHPLC columns packed with sub-2 micron particles.

HALO®BioClass UHPLC columns are specifically designed for fast, high-resolution separation of proteins and peptides. The particles packed in these columns are engineered to have pore sizes, particle sizes and diffusion path lengths that are optimum for the separation of proteins > 50 kDa (HALO Protein columns) and peptides in the range of 3 to 20 kDa (HALO Peptide columns).

The use of highly stable bonding chemistry ensures that separations are rugged and reliable and the low back pressure of these columns allow them to be used effectively on either UHPLC or conventional HPLC equipment.

HALO® BioClass Large Molecule Column Specifications

Bonded Phase	Pore Size (Å)	USP Designation	Particle Size (µ)	Carbon Load (%)	Surface Area (m²/g)	pH Range	Max.Temp Lower pH Limit	Max.Temp Upper pH Limit	Endcapped
Protein C4	1000	L26	2.7	0.6	22	2-9	90	40	Yes
Protein ES-C18	1000	L1	2.7	1.4	22	1-8	90	40	Yes
Protein C4	400	L26	3.4	0.4	15	2-9	90	40	Yes
Protein ES-C18	400	L1	3.4	1	15	1-8	90	40	Yes
Peptide ES-C18	160	L1	2 2.7 5	4.0 4.6 4.0	65 90 60	1-8	90	40	No
Peptide ES-CN	160	L10	2.7 5	2.2 1.5	90 60	1-8	90	40	Yes
Peptide Phenyl-Hexyl	160	L11	2.7	N.A	N.A	1-8	90	40	Yes
Glycan	90	L95	2.7	3.2	135	2-9	65	40	No

HALO Protein 1000Å

2.7 µm diameter spherical, ultra-pure, "Type B" silica
0.5 µm porous layer fused to a 1.7 µm solid core
1000Å pore size

HALO Protein 400Å

3.4 µm diameter spherical, ultra-pure, "Type B" silica
0.2 µm porous layer fused to a 3.0 µm solid core
400 Å pore size

Bonded Phase Available:

Protein C4: Dimethylbutylsilane, exhaustively endcapped and enhanced for high temperature stability.

ES-C18: Sterically-protected C18 phase

HALO Peptide

2.7µm (HALO) and 4.7µm (HALO-5) diameter spherical, ultra-pure, "Type B" silica
HALO: 0.5µm porous layer fused to a 1.7µm solid core
HALO-5: 0.6µm porous layer fused to a 3.5µm solid core
160Å pore size
Three bonded phases (ES-C18, Phenyl-Hexyl and ES-CN) provide alternate selectivity to optimize peptide separations.

Bonded Phases available:

Halo Peptide ES-C18: Octadecyldiisobutylsilane, Sterically-protected

Halo Peptide ES-CN: Cyanopropyldiisopropylsilane, Sterically-protected

Halo Peptide Phenyl-Hexyl: packed with 2.7-micron “core-shell” type particles having optimized pore sizes (160 Angstroms) and a third bonded phase (Phenyl-Hexyl) to complement the existing ES-C18 and ES-CN phases to provide more separation choices for peptide mapping.

Halo Glycan

2.7µm (HALO) diameter spherical, ultra-pure, "Type B" silica
HALO: 0.5µm porous layer fused to a 1.7µm solid core
90Å pore size

Bonded Phase

Incorporates a highly polar ligand that contains 5 hydroxyl groups tethered to 2.7µm Fused-Core silica particles via novel, proprietary linkage chemistry.

FOR SMALL MOLECULES PLEASE REFER TO HALO® 90Å

Guidance for Pore Size Selection

MW Range	Pore Size (Å)	HALO Column Family	HALO Product
< 5000 Da	90	HALO 90Å	C18, AQ-C18, C8, Phenyl-Hexyl, Biphenyl, PFP, ES-CN, RP-Amide, HILIC, Penta-HILIC
< 20 kDa*	90	HALO BioClass	HALO Glycan*
100 Da < MW < 15kDa	160	HALO BioClass	HALO Peptide (ES-C18, ES-CN, Phenyl-Hexyl)
2kDa < MW < 500kDa	400	HALO BioClass	HALO Protein (ES-C18, C4)
> 50kDa	1000	HALO BioClass	HALO Protein (ES-C18, C4)

*for glycans, glycopeptides and glycoproteins

Halo® UHPLC Guard Columns

Ultra-low dispersity for UHPLC columns
Effectively filters out particulate material and prevents column inlet frit plugging
Collects non-eluting compounds from the sample and prevents column fouling.
Finger-tight guard replacement design
Easily replace guard cartridge without removing guard holder from the flow path

Auto-adjusting ZDV fitting ensures optimum connection to the HALO analytical column

Simple adjustment to fit any UHPLC column

Halo® UHPLC guard columns are finger-tight, direct-connect units that represent the latest in fittings and column protection technology. They are easy to use, maintain ultra-low peak dispersion and are safe to operate at pressure up to 600 psi.

HALO C18 Care & Use
HALO C8 Care & Use
HALO HILIC Care & Use
HALO PFP Care & Use
HALO Phenyl-Hexyl Care & Use
HALO RP-Amide Care & Use