Flow Diagrams
Dr Ron Epping
LQB381 Unit Coordinator
1991 - 2012
EPPING, LQB381
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EPPING, LQB381
 Simple diagrams help you organise your mind and understand how
dilution factors apply to an experiment. Try working backwards
using AMOUNTS (g or moles) from the results of the test sample to
calculate the AMOUNT in the original sample.
Here is an example
100 uL of a protein extract is diluted 1/20 with saline and then assayed for
protein as follows:
250 μL of the test sample (diluted extract) is mixed with 100 μL of Reagent
A and 500 μL of Reagent B. The solution is then diluted to 3 mL with
d.water and the absorbance measured. From comparison with plotted
standards on a standard curve. the 3 mL sample is found to contain 1.5 mg
of protein.
What is the protein concentration of the original solution in mg/mL?
EPPING, LQB381
Flow Diagram: Example
Protein
PROTEIN
Solution
SOLUTION
1.0 mL
EPPING, LQB381
Flow Diagram: Example
0.1mL
Protein
PROTEIN
Solution
SOLUTION
0.1 mL Original Sample
1.0 mL
EPPING, LQB381
Flow Diagram: Example
0.1mL
Protein
PROTEIN
Solution
SOLUTION
1.9 mL saline
0.1 mL Original Sample
1.0 mL
EPPING, LQB381
Flow Diagram: Example
0.1mL
0.25 mL
Protein
PROTEIN
Solution
SOLUTION
1.9 mL saline
0.1 mL Original Sample
1.0 mL
2.0 mL
EPPING, LQB381
Flow Diagram: Example
0.1mL
0.25 mL
Protein
PROTEIN
Solution
SOLUTION
1.9 mL saline
0.1 mL Original Sample
1.0 mL
0.25 mL Test Sample
2.0 mL
EPPING, LQB381
Flow Diagram: Example
0.1mL
0.25 mL
0.25 mL
Protein
PROTEIN
Solution
SOLUTION
1.9 mL saline
0.25 mL Test Sample
0.1 mL Original Sample
1.0 mL
2.0 mL
0.25 mL
EPPING, LQB381
Flow Diagram: Example
0.1mL
0.25 mL
0.25 mL
Protein
PROTEIN
Solution
SOLUTION
1.9 mL saline
0.25 mL Test Sample
0.1 mL Original Sample
1.0 mL
2.0 mL
ABSORBANCE
1.5 mg
Read from the
standard curve
0.25 mL
ASSAY PROCEDURE
+ 0.1 mL A
+ 0.5 mL B
+ 0.15 mL H2O
1.0 mL
EPPING, LQB381
Flow Diagram: Example
0.1mL
0.25 mL
0.25 mL
PROTEIN
SOLUTION
1.9 mL saline
0.25 mL Test Sample
0.1 mL Original Sample
1.0 mL
2.0 mL
ABSORBANCE
1.5 mg
CALCULATE USING AMOUNTS:
1.5 x 10 -3 g x 2.0 / 0.25 x 1.0 / 0.1
= 0.12 g in 1 mL original sample
= 120 mg/mL
0.25 mL
ASSAY PROCEDURE
+ 0.1 mL A
+ 0.5 mL B
+ 0.15 mL H2O
1.0 mL
EPPING, LQB381
EPPING, LQB381