Hello Randa Elrayyes

Once you obtain the concentration of the unknown samples (diluted samples) from the standard curve, multiply that concentration by the dilution factor.

Suppose Sample A has a concentration of x ug/ml.

Sample A has 2 dilutions (1: 10 and 1:100).

Then the final protein concentration of sample A will be

10x ug/ml for 1:10 dilution and

100x ug/ml for 1:100 dilution.

.

Best.

@Malcolm Nober

Then get the sum of all results? That would be the whole concentration?

If you have obtained multiple measurements from the same sample tested at different dilutions, you can take the average. However, you should only use the measurements that fall within the range of the standard curve.

Hi Randa Elrayyes

No, you didn't get it right. Let me explain in detail.

As you have done three dilutions you need to find out which dilution has the absorbance in the linear range of the standard curve. The OD of only those dilutions which lie in the linear range of the curve will give the accurate protein concentration from the curve.

Let us suppose Sample A has three dilutions (1:10, 1:100 and 1:1000). If only 1:100 dilution has the absorbance in the linear range, then the protein concentration for that dilution will be most accurate.

If the protein concentration for 1:100 dilution is x ug/ml from the standard curve, then the final protein concentration will be 100x ug/ml after having multiplied by the dilution factor. This will be the final protein concentration or the whole concentration as you have mentioned and will correspond to the protein concentration of the stock.

Let us consider a second case.

If two dilutions lie in the linear range of the standard curve say 1:100 and 1:200 then after multiplying the protein concentration obtained from the standard curve by their respective dilution factors , you should take the average protein concentration. This will be the final protein concentration or the whole concentration which corresponds to the protein concentration of the stock.

I hope this is clearer now.

Good Luck.

Make a standard curve i.e. OD vs concentration with the help of different concentrations of protein (as you have done in different dilution). In excel you get a equation in the form of y=mx+c. m and c value is auto-calculated by excel. y is OD of unknown that you get my experiment. C is concentration that can be now calculated with the help of this equation.