I have a few questions:
What is an OD value?
Why do we use blank solution in Lowry's protein estimation method?
If The OD of a protein is 0.01, what does it mean?
Thanks in advance.
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1 Answer
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3
- The OD measurement is the output of what the photometer measures. It is actually the amount of light which is scattered or absorbed by your sample - scientifically called extinction.
- A blank is used to be able to substract the influence of reagents, light that is scattered on the surfaces of the cuvette (which is probably also not completely clean) and so on. Every of this influences reduces the amount of light which should reach the detector on the other side of the probe and give you false extinction readings. If you are using a colored detection sample (which colors deeper or looses color during the reaction) this is more obvious. The solution already has an extinction on its own which is not of interest. So you make a blank (which basically substracts all these influences from you sample reading) to correct for this. How this is done depends on the photometer. Some make a correction reading at the beginning, while others use a second cuvette (a blank cuvette) which is permanently measured.
- It means that the protein gives you this extinction. Without a calibration curve (made from serial dilutions with known protein concentrations) it is impossible to make any further comment on this number.
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$\begingroup$ Thanks a lot for quick response.but can you elaborate ans no. 2 please. $\endgroup$– N00BApr 28, 2014 at 16:43
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2$\begingroup$ You can think of the blank as a way of calibrating the instrument. The blank defines the amount of light that will hit the photodetector when there is none of the species of interest in the cuvette. By setting this as the zero you have prepared the instrument for measuring the samples of interest. $\endgroup$ Apr 28, 2014 at 18:18