Bernard, Ed,
Sounds like an impurity, all right. But if so you wold think
the chemical companies would be selling special low-A280 grade
imidazole for IMAC, and for some price you could get material
that doesn't absorb there at all. Unless its a spontaneous
decomposition product. Maybe even the pure compound has
some absorbance there, and impurities result in the
variability mentioned?
eab
Ed Pozharski wrote:
According to Qiagen
Since imidazole absorbs UV radiation at 280 nm, an elution profile
measured at 280 nm while purifying a 6xHis tagged protein by FPLC will
show an increase in absorbance above the background signal allowing
quantitation of your protein. The absorbance of imidazole can vary
depending on its source and purity, but elution buffer containing 250
mM imidazole usually has an A280 of 0.2–0.4.
Cheers,
Ed.
On Wed, 2013-08-21 at 11:25 -0400, Edward A. Berry wrote:
Would you happen to know what the A280 of a 1M (or .1 M) solution is?
I wonder if this absorbance is really due to impurities or is the tail of a
weak absorbance band of imidazole itself. I
notice A280 is not one of the properties sigma lists for its imidazole.
Jan wrote:
Hi Bernhard,
this one works for us:
Sigma BioUltra imidazole, >99.5% by GC, 56749
Cheers,
Jan
--
Jan Abendroth
Emerald BioStructures
Seattle / Bainbridge Island WA, USA
home: Jan.Abendroth_at_gmail.com
work: JAbendroth_at_embios.com
http://www.emeraldbiostructures.com
On Aug 21, 2013, at 7:33 AM, Bernhard Rupp <hofkristall...@gmail.com
<mailto:hofkristall...@gmail.com>> wrote:
Hi Fellows,
could someone please point me towards the source of a known high purity
imidazole
with low absorbance at 280 nm? I am facing the problem of detecting a low
absorption protein
in high imidazole background after IMAC gradient elution. In the UV spectra of
the
2 imidazoles I checked there is some contaminant that absorbs at 280…
Thx, BR
----------------------------------------------------------------------------------------
Bernhard Rupp
Marie Curie Incoming International Fellow
Innsbruck Medical University
Schöpfstrasse 41
A 6020 Innsbruck – Austria
+43 (676) 571-0536
bernhard.r...@i-med.ac.at <mailto:bernhard.r...@i-med.ac.at>
----------------------------------------------------------------------------------------
Dept. of Forensic Crystallography
k.-k. Hofkristallamt
Vista, CA 92084
001 (925) 209-7429
b...@ruppweb.org <mailto:b...@ruppweb.org>
b...@hofkristallamt.org <mailto:b...@hofkristallamt.org>
http://www.ruppweb.org/
-----------------------------------------------------------------------
--
Edwin Pozharski, PhD, Assistant Professor
University of Maryland, Baltimore
----------------------------------------------
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Then knowledge and wisdom are born along with hypocrisy.
When harmonious relationships dissolve then respect and devotion arise;
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------------------------------ / Lao Tse /
