quantitation of RNA

William R. Morgan wmorgan at ACS.WOOSTER.EDU
Sun Aug 15 15:21:24 EST 1993


>In article <00970F6E.227DD480.14784 at hal.hahnemann.edu>
>russos at HAL.HAHNEMANN.EDU writes:
>>HI,        I hope someone can help me.  I am isolating RNA from macrophages
>>           and consistently get such low amounts that quantitation by
>>           spectrophotometer is inaccurate.  Does anyone know a way to
>
>        The only thing that I can suggest is to use a smaller cuvette.  We
>are quite happy with a cuvette with listed volume of 50 ul, but I find that
>80 ul is required to fill the window on it.  I've seen Hellma advertising
>some cuvettes that only require 5 or 10 ul.  Alternatively, you can get
>a special type of small cuvette that requires a special small cuvette stage
>for Beckman spectrophotometers, but we're content not to have to keep
>changing the stage (or is it called the cuvette holder) within the spectro
>depending on who's using it.
>
>Mike

We've been quantitating small amounts of RNA using the Hellma
ultramicro-cuvette (minimal sample volume = 5 ul; typically we use 8 ul)
and a dedicated spectrophotometer.  Because the light path length is 5 mm,
you need an RNA conc. of 80 ug/ml for an 0D260 of 1.0.  So you can fairly
accurately measure as little as 40 ng of RNA (40 ng RNA in 5 ul volume =
OD260 of 0.1).

Bill Morgan

William R. Morgan
The College of Wooster
Department of Biology
931 College St.
Wooster, OH  44691
Phone:  216-263-2026
FAX:    216-263-2378
E-mail: wmorgan at acs.wooster.edu




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