Hydrophobic Agarose Beads

Hydrophobic interaction chromatography (HlC)has become a popular alternative to more classical affinity cromatography because of its ability to fractionate proteins using relatively less expensive ligands.

  • The use of hydrophobic dye ligands:
    because certain dyes mimic the hydrophobic structure of enzyme cofactors, they can be used to isolate the associated enzymes.
  • The use of aliphatic or phenyl ligands:
    linear, aliphatic molecules, when attached to a bead matrix, have the ability to intercalate into hydrophobic clefts on protein molecules, thereby slowing or stopping their movement through the column.
  • Advantages:
    the technique has the ability to remove unwanted, hydrophobic proteins like albumin (prior to 2D or further fractionation by other methods) or fractionate a heterogeneous population of proteins according to their relative hydrophobicity.
  • Disadvantages:
    multiple sites of attachment by some hydrophobic ligands can sometimes result in irreversible binding of protein to the ligand thereby reducing column utility. Or, the harsh conditions required to recover such tightly bound proteins can result in their denaturation.

Hydrophobic BioScience Beads:

Hydrophobic Micro-surfaces instead of small, linear ligands:

Hydrophobic BioScience Beads contain hydrophobic microparticles which allow a hydrophobic substance to bind to the surface of the microparticle instead of being trapped by ligand intercalation.

The very high porosity of the agarose beads allows most proteins to enter the bead and bind to the hydrophobic surfaces of the entrapped microparticles. 

The BioScience Bead design thereby reduces the likelihood of irreversible binding as well as increasing the amount of hydrophobic ligand mass on which to bind.

A wide range of hydrophobic microparticles can be made from one or more high M.P. waxes. The waxes can be divided into two categories, based on chemical composition:

  • High M.W. aliphatic or paraffin waxes:
    General formula : CH3- (CH2)n- CH3 , where n= 24 and the structure can contain linear, branched or alicyclic moieties.
  • High M.W. esterified waxes:
    General formula : R - COO - R' , where R and R' vary according to the type of wax. For example, R can represent fatty acids from C16 - C18 in beeswax to fatty acids from C16-C20 in Carnauba wax. Similarly, the R' alcohol can vary from C22-C28 alcohols in beeswax to C30-C34 alcohols in Carnauba wax.

The surface characteristics and binding properties of wax microparticles can be further modified with cationic, anionic, or neutral, hydrophilic moieties when they are micronized for inclusion in agarose beads. A wide range of surface characteristics can be achieved by varying the above parameters.

BioScience bead containing wax microparticles. 4% agarose bead; mag: 400X

Hydrophobic BioScience Beads Options

All Hydrophobic BioScience Beads are considered specialty beads and are made according to the option selections made by each customer. If you have not previously looked at the Standard & Specialty Bead Options page, it might be helpful to do so now before proceeding to the hydrophobic options listed below:

1 Agarose Concentration (%) Hydrophobic Particle Composition Incidence of Particles in Bead Surface Charge on Particles Bead Size Range Crosslinked Other Properties
2 2 High MW Aliphatic Wax High Neutral 50-150µ No None
3 4 High MW Esterified Wax Medium Anionic 250-350µ Yes Magnetic
4 6 Other: Specify Low Cationic Other: Specify Other: Specify
5 Other: Specify

Instructions for using Table of Options (above):
Select one property from each column.

Each column shows a set of options which can be combined with properties from other columns. Only one selection can be made from each column. The combined set of properties you select represent the specifications for your specialty, hydrophobic bead.


On the Order and Inquiry page, you need only enter the grid locations for each of the selections you've made, from the table above, in order to specify your hydrophobic bead properties. For example, one set of selections might be : A3 B3 C4 D3 E2 F2 G2 which would correspond to a 4% agarose bead containing high MW esterified wax micro-particles at a low incidence in the bead, each microparticle having an anionic surface charge. The bead would not be crosslinked or have any other special properties.


A price will be quoted to you when you make your inquiry. The price will be volume dependent but always attractive compared to any hydrophobic bead you're now using.

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Hydrophobic Beads Selection Guide

Note: bead size is 50-150 μ for all hydrophobic products; also, agarose concentration is either 4% (A-4B) or 6% (A- 6B) as indicated.

Reference Brand Name BioScience Bead Equivalent (code) Settled Bead Volume Catalog Number BioScience Bead Price Target Application(s)
Sigma A-3790; BSA-coupled Agarose Bead A-4B-CL-0R;
Albumin-binding, primarily hydrophobic, molecules
$96/5 ml 5 ml AW-A-0400-0005 $70.00
$172/10 ml 10 ml AW-A-0400-0010 $112.00
25 ml AW-A-0400-0025 $224.00
100 ml AW-A-0400-0100 $716.00
> 100 ml Inquire Inquire
No commercial counterpart A-4B-µwax-anionic 10 ml AW-A-0400-0010 $25.00 Cationic and/or hydrophobic proteins or lipids
50 ml AW-A-0400-0050 $100.00
100 ml AW-A-0400-0100 $160.00
200 ml AW-A-0400-0200 $256.00
400 AW-A-0400-0100 $410.00
> 400 ml Inquire Inquire
No commercial counterpart A-4B-µwax-cationic 10 ml AW-A-0400-0010 $25.00 Anionic and/or hydrophobic proteins, nucleic acids or lipids
50 ml AW-A-0400-0050 $100.00
100 ml AW-A-0400-0100 $160.00
200 ml AW-A-0400-0200 $256.00
400 AW-A-0400-0400 $410.00
> 400 ml Inquire Inquire
No commercial counterpart A-4B-µwax-neutral 10 ml AW-A-0400-0010 $25.00 Uncharged, hydrophobic proteins or lipids
50 ml AW-A-0400-0050 $100.00
100 ml AW-A-0400-0100 $160.00
200 ml AW-A-0400-0200 $256.00
400 AW-A-0400-0400 $410.00
> 400 ml Inquire Inquire
No commercial counterpart A-6B-µwax-anionic 10 ml AW-A-0600-0010 $25.00
50 ml AW-A-0600-0050 $100.00
100 ml AW-A-0600-0100 $160.00
200 ml AW-A-0600-0200 $256.00
400 AW-A-0600-0400 $410.00
> 400 ml Inquire Inquire
No commercial counterpart A-6B-µwax-cationic 10 ml CW-A-0600-0010 $25.00 Anionic and/or hydrophobic proteins, nucleic acids or lipids
50 ml CW-A-0600-0050 $100.00
100 ml CW-A-0600-0100 $160.00
200 ml CW-A-0600-0200 $256.00
400 CW-A-0600-0400 $410.00
> 400 ml Inquire Inquire
No commercial counterpart A-6B-µwax-neutral 10 ml NW-A-0600-0010 $25.00 Uncharged, hydrophobic proteins or lipids
50 ml NW-A-0600-0050 $100.00
100 ml NW-A-0600-0100 $160.00
200 ml NW-A-0600-0200 $256.00
400 NW-A-0600-0400 $410.00
> 400 ml Inquire Inquire

A=Agarose; 4 or 6 = % Agarose in gel bead; CL=crosslinked; R= ether-linked substituent or derivative; μwax = micro-wax microspheres; AW = anionic wax; CW = cationic wax; NW = neutral wax

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