I am cultivating a culture of anaerobic archaea. The growing medium includes sodium resazurin as anoxic indicator. I would like to estimate the cell concentration of the culture by measuring the absorbance of the sample with UV vis spectrophotometer. However, I encounter two issues with this measurement.
1) As I withdraw a sample from my serum bottle and place it into a cuvette, the sample slowly changes colour since the environment is no longer anoxic. This makes the absorbance reading constantly change (due to the gradual change in colour of the indicator).
2) Will the cells sink down to the bottle of the cuvette, thus make my measurement inaccurate?
Use a UV-Vis well plate reader instead of using a cuvette mounted normal uv-vis system to eliminate the precipitation problem.
you may use a closed sample cell, fully filled. As measurement can be conducted in seconds error by settling should be tolerable, us as high as possible wavelength to minimize effect of colour.
You can overcome the problem of oxygen as Titus suggested. Many spectrometers are equipped with a magnetic stirrer under the cuvette holder to prevent sedimentation by continuous stirring.
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- Methodology
- Absorption Spectroscopy
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