Automated assay for determining cellular cholesterol using a random access chemistry analyser.

BACKGROUND

We have previously reported an ultrasensitive fluorometric assay for measuring cellular cholesterol. Although this technique is reliable, the use of the assay has limitations due to the requirement for special equipment. It is therefore difficult to apply this assay for the routine determination of cellular cholesterol.

METHODS

A colorimetric assay to measure cellular cholesterol was established that utilizes reagents widely used for the measurement of cholesterol in blood samples in conjunction with a random access chemistry analyser ARCHITECT c8000 that is also common in clinical laboratories.

RESULTS

This colorimetric assay showed excellent linearity and recovery. The within-run coefficients of variation were less than 2.5%. The sensitivity of this method, with its detection limit of 1.29 mumol/L, was found to be superior to that of the fluorometric assay we have developed previously. In platelets obtained from patients with diabetes, both the free cholesterol and cholesterol ester content were significantly increased.

CONCLUSIONS

Using this technique, measurement of cellular cholesterol could be performed routinely without the requirement for special reagents and equipment.