Article View: sci.techniques.spectroscopy
Article #22Re: Drug binding: When fluorescence polarization fails
From: Marvin
Date: Tue, 02 Oct 2007 15:50
Date: Tue, 02 Oct 2007 15:50
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3124 bytes
Dominic-Luc Webb wrote: > Thanks Marvin for your response, and sorry about any confusion I > might have created in the use of the word "strangely". The > "strangeness" had only to do with the disjunct between the > unexpected glycerol data and the expected protein binding data > using the same labelled drug. Until now, I regarded both of > these as positive control tests. > > Regarding 50% glycerol, I could be convinced that the fluorescence > lifetime was changed, but I would need to bring this in line with > recent experiments. I went back and made some further measurements > and quantifications together with data I collected earlier. I > calculate a 1.4 fold increase in fluorescence intensity (FI) when > measured with normal FI filter set. Using the FP filters to measure > same samples, there was also an increase in fluorescence intensity in > both of the two channels (1.35 in the static channel, 1.41 in the > perpendicular channel). Fluorescence yield must have increased > concommitantly with a differential shift towards the perpendicular > channel. > > Considering that last sentence, this assay has thus far not been > very sensitive to different intensities (e.g., fluorescein over a > few orders of magnitude). However, the differential shift towards > the perpendicular channel (1.41/1.35), although small, approaches > significance and seems in line with your suggestion that fluorescence > lifetime was decreased. Does this make sense? > > Dominic When you change the environment around a molecule, energy levels change. Try scanning the absorption and emission spectra, which should be different in 50 glycerol vs. water. It isn't easy to generalize about specific changes in optical properties.
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X-Google-Thread: fe222,d7292fc16d0f3513 X-Google-Thread: fd9c4,d7292fc16d0f3513 X-Google-Attributes: gidfe222,gidfd9c4,public,usenet X-Google-Language: ENGLISH,ASCII-7-bit Path: arc1-ca.pugleaf.net!g2news2.google.com!news1.google.com!border1.nntp.dca.giganews.com!border2.nntp.dca.giganews.com!nntp.giganews.com!cyclone1.gnilink.net!spamkiller2.gnilink.net!gnilink.net!trndny03.POSTED!9fcca4b5!not-for-mail From: Marvin <physchem@verizon.net> User-Agent: Thunderbird 2.0.0.6 (Windows/20070728) MIME-Version: 1.0 Newsgroups: sci.techniques.spectroscopy,bionet.molbio.proteins Subject: Re: Drug binding: When fluorescence polarization fails References: <Pine.LNX.4.44.0710011540080.19573-100000@uno.canit.se> <049Mi.11183$bV2.6674@trndny02> <Pine.LNX.4.44.0710021021310.3383-100000@uno.canit.se> In-Reply-To: <Pine.LNX.4.44.0710021021310.3383-100000@uno.canit.se> Content-Type: text/plain; charset=ISO-8859-1; format=flowed Content-Transfer-Encoding: 7bit X-Antivirus: avast! (VPS 000778-1, 10/02/2007), Outbound message X-Antivirus-Status: Clean Lines: 33 Message-ID: <7NtMi.4879$Jk2.3800@trndny03> Date: Tue, 02 Oct 2007 15:50:27 GMT NNTP-Posting-Host: 71.167.164.128 X-Complaints-To: abuse@verizon.net X-Trace: trndny03 1191340227 71.167.164.128 (Tue, 02 Oct 2007 11:50:27 EDT) NNTP-Posting-Date: Tue, 02 Oct 2007 11:50:27 EDT X-Original-Bytes: 2941