Question

Calculate the volume needed to produce the standard curve in µl

Part I: Standard Curve 1. We will use your 25 mMD-glucose solution to make a standard curvel Conceptually, this is very similar to the Bradford standard curve that you-have done before. The goal is to define the mathematical relationship between absorbance-and amount of teducing,carbohydrate in the reaction.

Answer 1

Stock glucose solution: 25 mM or 25 mmole/L or 25,000 µmoles/L

To get working concentration of 7.5 µmoles following aliquote is needed:

(think like if 1000 mL for 25,000 µmoles/L then how much aliquote for 7.5 µmoles)

i.e

$\frac{1000}{25,000}\times 7.5$

= 0.3 mL or 300 µl

similarly, for 5 µmoles

$\frac{1000}{25,000}\times 5$

= 0.2 mL or 200 µl

for 2.5 µmoles,

$\frac{1000}{25,000}\times 2.5$

= 0.1 mL or 100 µl

for 1.25 µmoles,

$\frac{1000}{25,000}\times 1.25$

= 0.05 mL or 50 µl

for 0.5 µmoles,

$\frac{1000}{25,000}\times 0.5$

0.02 mL or 20 µl

for 0.25 µmoles,

$\frac{1000}{25,000}\times 0.25$

= 0.01 mL or 10 µl.

Working concentration of glucose (µmoles)	Aliqute from stock solution (µL)	D/W to make up volume to 300 µL	DNS solution (µL)	Incubate at 75 º C for 15 minutes	Tartrate solution (µL)	A570nm
0	0	300	500		200
0.25	10	290
0.5	20	280
1.25	50	250
2.5	100	200
5	200	100
7.5	300	0