Alex's insights not withstanding, Milton Hearn published an article a
number of years ago in J. Chromatography in which he showed that loadings
on non-porous columns could be equivalent to fully porous packings, since
ligand density and in this case, accessible ligand, are the driving factors
for load. This is especially true for ion exchange of DNA.
The work that I have done in conjunction with NCI researchers on P3 and PUC
19 separations showed that the porous medias were not capable of
differentiating between the large DNA species. Use of non-porous medias
gave a consistent surface presentation to the molecules, and thus better
resolution. We attributed this to lack of consistent accessibility of the
surface ligands because of molecular exclusion from the "small (900-1500A)
pores."
Additional data can be gleaned from the Macherey-Nagel Nucleogen product
series, where the 500A column is better suited for the digests and the
4000A product for intact DNA. However, all of these silica based medias
suffer from the same limitation: poor resistance to high pH elution
requirements of certain DNA types. Consequently, a polymer based ion
exchanger, while having lower pore volume, hence lower capacity from that
aspect, are far more robust for a wider range of applications, especially
if they have a higher ligand density to compensate for pore volume
limitations.
The Biochrom QA-NP10 or NS-NP10, the TSK DEAE-NPR and the Waters GenPak
columns are all polymeric offerings. The latter two are 2.5 micron
products which reuire some small volume capapbility from your HPLC systems
to capture their resolving power.
Amos Heckenodrf , Ph.D.
The Nest Group, Inc.
Compare our NEW, SuproTipÅ family of coated wall chemistries to ZipTips.
See: http://world.std.com/~nestgrp/protocols/AmiKa/SuproTip.pdf for more
information.
Value Added Resellers of HPLC Columns (VydacÅ, PolyLCÅ, BioChrom
HydrocellÅ, Jordi-GelÅ, Macherey-Nagel NucleogenÅ, Higgins Analytical
HAISilÅ100,TARGAÅ & CLIPEUSÅ, and Bischoff ProntosilÅ) Genomic SolutionsÅ
PAGE gels; Amika Sialomed COZAPÅ destaining pads, BioDialyserÅ,
Dispo-BioDialyserÅ and SuproTipÅ MS MicroSample SPE Tips; and KronLab
BioCartÅ and PROÅ glass adjustable columns.
Tel: 800-347-6378 or 508-481-6223; FAX 508-485-5736;
The Nest Group, Inc., 45 Valley Rd, Southboro, MA 01772-1306
>Hi Gordon,
>
>I'd like to point out that non-porous materials aren't the only option. We
>have demonstrated HPLC as both sizing and DHPLC applications on a Wide Pore
>Silica based material. Porous materials have the advantage of high sample
>capacity for semipreparative work. This is marketted by HP as Eclipse
>dsDNA. LEt me know if you have any other questions.
>
>Alex Apffel, Ph.D.
>Biomeasurements Group
>Hewlett-Packard Laboratories
>3500 Deer Creek Road
>Palo Alto, CA 94304
>Tel: 650-857-6090
>Fax: 650-852-8502
>Email: alex_apffel@hpl.hp.com
>
>
>-----Original Message-----
>From: Gordon Alton [mailto:galton@signalpharm.com]
>Sent: Friday, August 06, 1999 7:08 PM
>To: Recipients of ABRF List
>Subject: DNA/RNA chromatography
>
>
>Dear ABRFers,
>
>
>Does anyone have experience with DNA or RNA chromatography on non-porous
>reverse phase resin? Which chromatography companies have this type of resin?
>I would greatly appreciate any information. In advance, thanks.
>
>
>--------------------------------------------------------
>Gordon Alton, PhD
>
>Gene and Protein Discovery Group
>
>Signal Pharmaceuticals Inc.
>5555 Oberlin Drive
>San Diego, CA 92121
>
>Email: galton@signalpharm.com
>Phone: 619-558-7500 x8252
>Fax: 619-623-0870
>WWW: http://www.signalpharm.com
>-------------------------------------------------