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bacterial OD

bacterial OD - Microbiology Forum

bacterial OD - Discuss Microbiology Science and Protocols here. Post questions on the study of viruses, fungi, parasites and bacteria here. Microbiology Forum.


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  #1  
Old 10-15-2003, 08:18 AM
Vestitas
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Default bacterial OD



i've noticed that most bacterial ODs are measured at 600nm this being a
wavelength at which the light is scattered and not absorbed by the cells (?)
therefore the higher the OD the more turbid the culture. So why do some people
measure OD at 660nm? - Please help, I'm confused
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  #2  
Old 10-15-2003, 11:22 AM
Lesley Robertson
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Default bacterial OD


"Vestitas" <[Only registered users see links. ]> wrote in message
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people

We always did it at 430. It doesn't really matter what you use as long as
you standardize everything, use a good blank and make sure that you're
measuring over a range where your OD meter is linear.
Lesley Robertson
[Only registered users see links. ]


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  #3  
Old 10-15-2003, 03:04 PM
Vestitas
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Default bacterial OD

Cheers - v. helpful!
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  #4  
Old 10-15-2003, 06:15 PM
EK
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Default bacterial OD


"Vestitas" <[Only registered users see links. ]> wrote in message
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It does not generally matter for E.coli, 550, 600, or 660 nm. Cells of this
bacterium, as well as of most other bacteria, do not absorb much in this
area. However, one shouold be aware that overexpression of some proteins
might affect absorption spectra of the culture. It is always better checking
the 400-800nm spectrum of the particular culture you are using and selecting
the wavelength in the "pit" area where the absorbance stays unchanged in at
least the +-20nm range.
-Emir


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  #5  
Old 10-15-2003, 06:53 PM
DKafkewitz
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Default bacterial OD

Light scattering varies inversely with wavelength. The shorter the wave the
more the scatter. Hence 440 is more sensitive than 660. But absorbance of the
light by the medium is also important. Colorless media abosrb no visible
light: that is why they are colorless. Yellow media absorb in the shorter
range. Therefore you use a longer wave to minimize the false reading of
Absorbance rather than scattering.


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  #6  
Old 10-15-2003, 08:49 PM
EK
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Default bacterial OD


"DKafkewitz" <[Only registered users see links. ]> wrote in message
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the
the
BTW, I always wondered would then it be necessary to use shorter wavelengths
for smaller bacteria? I guess a 300nm monococci would not scatter light at
600nm, whereas 300nm diplococci would. Same would the rod e.g. 200 x 1000
nm. Is my understanding correct?
Emir


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  #7  
Old 10-16-2003, 08:05 AM
Lesley Robertson
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Default bacterial OD


"DKafkewitz" <[Only registered users see links. ]> wrote in message
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the
the
If you set zero on a sterile blank of the same medium, or use a double beam
with a sterile blank, you cancel the effect of the medium unless the culture
is producing a pigment. We always used 430nm with our bacteria because it
gave a longer linear area of ODs with our spectometers, cutting out the
amount of dilution needed. One common mistake I've seen students make is to
dilute with water rather than sterile medium, thereby introducing another
variable.
Lesley Robertson
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  #8  
Old 10-16-2003, 06:53 PM
Des O'Connor
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Default bacterial OD


"Lesley Robertson" <[Only registered users see links. ]> wrote in message
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of
visible
shorter
beam
culture
to

Also you could try a Nephalometer (Nephalometry)


Best Des



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  #9  
Old 10-17-2003, 05:38 AM
EK
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Default bacterial OD

> Also you could try a Nephalometer (Nephalometry)
It's nephelometry, from Greek 'nephel' for cloud (= meter of cloudiness).


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  #10  
Old 10-17-2003, 03:44 PM
Des O'Connor
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Default bacterial OD

Thanks for the correction and I thought spelling was my only strong point
lol

D

"EK" <[Only registered users see links. ]> wrote in message
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