Plant-tc Monthly Archive - December 1998

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Re: Lowry assay vs Bradford assay



Dear Claude,

The lipids shouldn't be a problem if you sample from below the lipid layer
immediately after centrifuging.   If I'm remembering right, SDS interferes
with even the modified Lowry assay except at very low SDS concentrations.
Maybe that's your problem.  The Bradford assay definitely doesn't work with
SDS.  Nor do any of the fluorescent assays I've tried.   The BCA protein
assay kit sold by Pierce is the best I've found for use with buffers
containing SDS.   Best of luck to you.

dr. dave


David E. Culley  Ph.D.  AMSD
Glass Garden Research
745 NW Darrow St.
Pullman, WA 99163

(509) 334-4884
dculley@pullman.com

-----Original Message-----
From: CLAUDE YVES BOMAL <abj206@AGORA.ULAVAL.CA>
Date: Tuesday, December 01, 1998 12:48 PM
Subject: Lowry assay vs Bradford assay


>Hello, I got some problems in protein concentration determination from
>somatic embryo extract. Actually, I use the bradford assay kit (Biorad)
>for soluble protein assay: It works quite well with very good
>reproductibility. Due to the composition of the extraction buffer with
>insoluble proteins, we opted for Lowry assay (DC). However, the
>OD is  extremely variable from one measurement to another: for the same
>extract, the OD (655nm) ranged from 0.612 to 0.917. The extraction buffer
>is 65 mM Tris pH6.8, 2% SDS, 10% Glycerol (Misra and Green, 1990).
>
>What is (are) the problem(s)?
>
>Shall I use the bradford assay kit for the insolubles protein extract?
>What about SDS interference in that case?
>
>I also noticed the presence of lipids at the surface of extracts after
>centrifugation. Is it a problem?
>
>All advices will be welcome,
>
>Greatings Claude.
>
>Claude Bomal
>Centre de recherche en biologie forestiere
>Pavillon Marchand, Laval University,
>Quebec, Qc, G1S 7P4, Canada
>email: abj206@agora.ulaval.ca

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