Cholesterol assay

Question

1 ml of whole blood is collected and centrifuged to obtain a plasma sample. The volume of the plasma sample is 300 microlitres. A 1 in 4 dilution of the plasma is made and analysed using a cholesterol assay like the one you performed in the laboratory. 10 microlitres of both the diluted plasma sample and a cholesterol standard are assayed using 1mL of cholesterol reagent. Following incubation at 37 degrees Celcius, the samples are measured spectrophotometrically at 500nm.

If the absorbance of the plasma sample assay was 0.5, the absorbance of the cholesterol standard assay was 0.4, and the concentration of cholesterol in the standard was 1.3mM, what was the concentration of the cholesterol in the whole blood (in mM)?

 

(assume all of the cholesterol in the whole blood sample was recovered in the plasma; do not include units in your answer; give your answer to 2 decimal places)

Answer 1

Step 1:

Volume of the plasma sample is 300 microlitres.

Calclulate the number of the moles of cholesterol in the plasma sample.

Number of moles can be calculated using formula , where c is the concentartion and v is the volume.

Calculate n.

Substitute and .

Therefore, .

Concentration of the cholesterol in the whole blood is 4.33 mM.

Step 2:

Calclulate the concentration of the cholesterol in the whole blood.

.

Substitute , and .

This concentration,   is the diluted concentration.

Calculate the undiluted concentration using the dilution factor, which in this case is 4.

Concentration of the cholesterol in the whole blood is 21.65 mM.

Solution:

Concentration of the cholesterol in the whole blood is 21.65 mM.