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<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'>Using DNA absorbance is exactly what we would do/have done if: <o:p></o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'><o:p> </o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'>We knew the DNA payload/phage. <o:p></o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'>We were not certain of the particle to pfu (plaque forming unit)
ratio, or wanted particles not infectious particles<o:p></o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'>We could dissociate the phage (usually we use 6 M GuHCl and some
heating if necessary) to eliminate scattering<o:p></o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'>The protein to DNA ratio was <= 1 (a good approximation for
spherical phage) to minimize protein contribution<o:p></o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'>We could not grow them on a host(s). <o:p></o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'><o:p> </o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'>Far and away if you can grow them tittering is the easiest way
to get concentration of infectious units. Lower limit (without concentration)
is about 10E2-3 particles/ml and upper limit is essentially infinite. <o:p></o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'><o:p> </o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'>Since phage are the most abundant organism on earth (10E7/ml
seawater <a href="http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1110918/">http://www.ncbi.nlm.nih.gov/pmc/articles/PMC1110918/</a>)
it is important to keep track of them. </span><span
style='font-size:11.0pt;font-family:Wingdings;color:#1F497D'>J</span><span
style='font-size:11.0pt;font-family:"Calibri","sans-serif";color:#1F497D'><o:p></o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'><o:p> </o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'>Did anybody take a look at the nanosight ( <a
href="http://www.nanosight.com/">http://www.nanosight.com/</a>) I
mentioned earlier? It is a pretty clever instrument that would seem to be
perfect for this application. Somewhat like DLS except individual particle
tracking makes it more suitable for polydisperse mixtures. We were able to
resolve and quantify a mixture of T=4 and T=7 capsids, detect the faction of
particles with and without tails, etc. And it is sort of based on hydrodynamics
so it fits the original request. <o:p></o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'><o:p> </o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'>Peter<o:p></o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'><o:p> </o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'><o:p> </o:p></span></p>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'><o:p> </o:p></span></p>
<div>
<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'>Peter E. Prevelige Jr.<br>
Professor<br>
Dept. of Microbiology, BBRB 416/6<br>
Univ. of Alabama @ Birmingham<br>
845 19th St. South<br>
Birmingham AL. 35294-2170<br>
Phone 205 975-5327<br>
FAX 205 975-5479<br>
<a href="mailto:prevelig@uab.edu">prevelig@uab.edu</a><br>
<a href="http://www.microbio.uab.edu/faculty/prevelige/prevelige-p.htm">http://www.microbio.uab.edu/faculty/prevelige/prevelige-p.htm</a><o:p></o:p></span></p>
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<p class=MsoNormal><span style='font-size:11.0pt;font-family:"Calibri","sans-serif";
color:#1F497D'><o:p> </o:p></span></p>
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<div style='border:none;border-top:solid #B5C4DF 1.0pt;padding:3.0pt 0in 0in 0in'>
<p class=MsoNormal><b><span style='font-size:10.0pt;font-family:"Tahoma","sans-serif"'>From:</span></b><span
style='font-size:10.0pt;font-family:"Tahoma","sans-serif"'>
rasmb-bounces@rasmb.bbri.org [mailto:rasmb-bounces@rasmb.bbri.org] <b>On Behalf
Of </b>Ariel Lustig<br>
<b>Sent:</b> Tuesday, March 09, 2010 3:33 AM<br>
<b>To:</b> rasmb<br>
<b>Subject:</b> [RASMB] phage con<o:p></o:p></span></p>
</div>
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<p class=MsoNormal><o:p> </o:p></p>
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<p class=MsoNormal><span style='font-size:10.0pt;font-family:"Arial","sans-serif"'>Dear
all,</span><o:p></o:p></p>
</div>
<div>
<p class=MsoNormal><span style='font-size:10.0pt;font-family:"Arial","sans-serif"'>I
just looked in the<u> Biophysical Chemesrty</u> 59 paper 1996 41 - 59 E
Kellenberger , E Staufer, M Haener , A Lustig, D.Karamater, >> Mechanism
of long tail-fiber devel. of bacteriophages.......<<
we determined the phage concentration with
UV Absorption 280nm.....ariel</span><o:p></o:p></p>
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<p class=MsoNormal><span style='font-size:10.0pt;font-family:"Arial","sans-serif"'><a
href="mailto:ariel.lustig@bluewin.ch">ariel.lustig@bluewin.ch</a></span><o:p></o:p></p>
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