[RASMB] determination of phage particle concentration

Mon Mar 8 07:44:23 PST 2010

Hi-
The wavelength dependence will differ from 4 due to dispersion.
A couple of papers to consider:
Camerini-Otero, R. D., and L. A. Day. 1978. The wavelength dependence
of the turbidity of solutions of macromolecules. Biopolymers. 17:
2241-2249.
Camerini-Otero, R. D., R. M. Franklin, and L. A. Day. 1974. Molecular
weights, dispersion of refractive index increments, and dimensions from
transmittance spectrophotometry. Bacteriophages R17, T7, and PM2,
and tobacco mosaic virus. Biochemistry. 13:3763-3773.
Best wishes,
Tom

Glen Ramsay wrote:
> Greetings RASMB:
>
> The scattering increases as the fourth power of the inverse 
> wavelength.  Years ago Aviv's spectrophotometer had a function that 
> would fit the baselines before and after an absorbance peak to 
> subtract the scattering portion. 
>
> However, I cringe at the idea of making analytical measurements of 
> particles via absorbance.  Absorbance of a photo can happen only once. 
> Scattering can happen more than once, sending a photon back into the 
> detector's view.  The detectors of different instruments view 
> different collection angles of the sample, so the amount of scattered 
> light is going to vary depending on the instrument design.  
> Reflections of scattered light off the surroundings can send photons 
> back onto the detector. In one custom instrument Aviv mounted the 
> detector on a rail, and apertures were mounted on both the sample's 
> exit and detector's entrance, to control the light collection angle.  
> And of course the size of the particle affects the amount of light 
> scattering.
>
> Measure scattering with absorbance with both eyes and mind open, 
> because the instrument is not being used as intended.
>
> Glen
>
>
> At 09:44 AM 3/5/2010, Steve Harding wrote:
>> Content-class: urn:content-classes:message
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>>          boundary="----_=_NextPart_001_01CABC72.57041806"
>>
>> Dear Sabine
>> Simple turbidity measurements (making sure you are _away_ from 
>> absorption maxima) using a good quality spectrophotometer _may 
>> _suffice for the sort of information you are after - particle 
>> concentration, although stricvtly speaking it is not a hydrodynamic 
>> method..  This is is the usual method for measuring concentrations 
>> (particles per ml) of very large assemblies such as spores and I 
>> think it works for smaller assemblies that are not too dilute..
>> Victor Bloomfield's group did quite a bit on the turbidity of phages 
>> in the late 70's and gave the relevant formulae (you may need the 
>> approx  mol. wt value) - if you look up Bahls and Bloomfield (and not 
>> Victor's QLS papers) that should give you a lead!
>> All best
>> Steve Harding
>>  
>> / http://www.nottingham.ac.uk/ncmh
>> /
>> > Dear all,
>> >
>> > we are trying to obtain the concentration of bacteriophages in a
>> > solution, concentration meaning the number of particles per ml. The
>> > solution is supposed to be monodisperse and the MW is (roughly) known
>> > but could be determined exactly. Is the any idea of how to do this
>> > with a hydrodynamic method (staining is difficult to compare between
>> > phage mutants and counting infected cells is tedious). I`ve got the
>> > "feeling" that one could use the number-averaged MW, but maybe this is
>> > totally wrong.
>> >
>> > Thank you for any hints, literature is also welcome.
>> >
>> > Cheers,
>> >
>> > Sabine
>> >
>> >
>> >
>> >
>> > Sabine Kaltofen
>> > PhD student
>> >
>> > Universität Potsdam
>> > Department of Physical Biochemistry
>> > Institute of Biochemistry and Biology
>> > Karl-Liebknecht-Str. 24-25, Haus 25, Raum B/0.05
>> > D-14476 Potsdam-Golm
>> > Telefon: +49-(331)-977-5245
>> > Email: kaltofen at uni-potsdam.de
>> >
>> >
>>
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>
>    Glen Ramsay, Ph.D.
>    Chief Scientist
>    Aviv Biomedical, Inc.
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