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Measurement of adrenal corticosteroids by high-performance liquid chromatography
1Department of Medical Technology, Bowling Green State University, Ohio 43403.
Objective:
To develop a high-performance liquid chromatography (HPLC) method for simultaneous measurement of adrenal corticosteroids important for the diagnosis of congenital adrenal hyperplasia.
Design:
Blind comparison using convenience samples.
Setting:
Research laboratory at the Department of Medical Technology, Bowling Green State University.
Patients:
Referred samples for cortisol, 11-deoxycortisol (S), and 17-hydroxyprogesterone (OHP). Stimulation condition unknown.
Intervention:
Venous blood collected, processed to serum, and frozen. Adrenal steroids measured by radioimmunoassay (RIA) and by HPLC.
Main Outcome Measures:
Linearity, analytic sensitivity, accuracy, and precision were evaluated using recovery, coefficient of variation (c.v.), correlation coefficient (r), Student's t test, F test, and linear regression.
Results:
Calibration curves were linear to at least 50 mumol/L, and the low-end sensitivity was 10 nmol/L. Analytic recovery ranged from 89.8% to 103.9%. The CV was below 7.5% (n = 10) for all three steroids. Comparison to RIA yielded r values of 0.909 (n = 25), 0.932 (n = 10), and 0.983 (n = 10) for cortisol, S, and OHP, respectively. Student's t-test results were 1.13 (p = 0.27), 0.10 (p = 0.93), and 0.48 (p = 0.64) for cortisol, S, and OHP, respectively. F-test results were 1.14 (p = 0.75), 1.37 (p = 0.66), 2.39 (p = 0.21) for cortisol, S, and OHP, respectively. Results for cortisol showed a negative bias of 97.0 nmol/L (3 mug/dL). Linear regression analysis revealed proportional errors of -15.0% -7.0%, and +9.0%, for cortisol, S, and OHP, respectively.
Conclusion:
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