Hi Jeroen,
We just bought a Stratagene Mx3005p for the Thermofluor method (also
known as differential scanning fluorimetry). This was after talking to
Martin, among others, he he... We haven't had it long and did our
first experiments last Friday, but it produced good results straight
away. We chose the Stratagene for a couple of reasons:
a) the iQ5 required a manual change of filter which would make it
unuseable for qPCR, and while we don't anticipate a great deal of
usage for qPCR it would be a shame to cripple such an expensive
machine. The filter for the Mx3005p can be chosen in software.
b) the Mx3005p uses a photomultiplier instead of a CCD to read the
plate. This (according to Stratagene) leads to better uniformity of
detection across the plate. I believe the iQ5 requires some kind of
calibration run each time you use it.
Our first impression of the Stratagene software is very positive. We
haven't tried the iQ5 but others say it's a perfectly fine machine. A
couple of people who know more than most are Helena Berglund at SGC
Stockholm (BioRad) and Frank Niesen at SGC Oxford (Stratagene).
Whichever of these two machines you choose, SGC have produced an
excellent detailed protocol and from their FTP site ftp.sgc.ox.ac.uk
you can download Excel spreadsheets to calculate Tm and produce at-a-
glance output for up to 96 wells from either iQ5 or Mx3005p output.
SGC intend to continue supporting output from both machines.
There are cheaper models by BioRad which we didn't look into (we got a
grant for the Mx3005p!) but I imagine they have sensitivity/wavelength
issues.
The link to the SGC protocol is:
http://www.ncbi.nlm.nih.gov/pubmed/17853878?ordinalpos=3&itool=EntrezSystem2.PEntrez.Pubmed.Pubmed_ResultsPanel.Pubmed_RVDocSum
HTH
Derek
On Feb 12, 2008, at 18:07, mesters wrote:
Sorry for the off-topic but can somebody recommend highly a
sensitive RT-PCR machine for the thermofluor experiment (sypro
orange).
That would imply excitation below 500 nm (ideally 470) and detection
at about 570 nm, right? I know several simple machines have a
problem with the 570 nm...
Jeroen.
--
Jeroen Raymundus Mesters, Ph.D.
Institut fuer Biochemie, Universitaet zu Luebeck
Zentrum fuer Medizinische Struktur und Zellbiologie
Ratzeburger Allee 160, D-23538 Luebeck
Tel: +49-451-5004070, Fax: +49-451-5004068
E-mail: [EMAIL PROTECTED]
Http://www.biochem.uni-luebeck.de
Http://www.iobcr.org
Http://www.opticryst.org
--
If you can look into the seeds of time and say
which grain will grow and which will not - speak then to me (Macbeth)
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