DBC Aldosterone ELISA

PRINCIPLE OF THE TEST 
The principle of the following enzyme immunoassay test follows the typical competitive binding scenario. Competition occurs between an unlabelled antigen (present in standards, controls and patient samples) and an enzyme-labelled antigen (conjugate) for a limited number of antibody binding sites on the microplate wells. The washing and decanting procedures remove unbound materials. After the washing step, the enzyme substrate is added. The enzymatic reaction is terminated by addition of the stopping solution. The absorbance is measured with a microplate reader. The intensity of the colour formed is inversely proportional to the concentration of aldosterone in the sample. A set of standards is used to plot a standard curve from which the amount of aldosterone in patient samples and controls can be directly read.

CLINICAL APPLICATIONS 
Aldosterone is a potent mineralocorticoid whose synthesis and release are controlled by the renin-angiotensin system of the body. Aldosterone promotes the reabsorption of sodium in the distal tubules of the kidney resulting in potassium secretion along with sodium retention, which controls the circulating blood volume. Chronic overproduction and secretion of aldosterone leads to hypertension. 
Measurement of aldosterone levels in serum in conjunction with plasma renin activity levels can be used to differentiate between primary and secondary aldosteronism.

SPECIMEN COLLECTION AND STORAGE 
Serum: Approximately 0.2 mL of serum is required per duplicate determination. Collect 4–5 mL of blood into an appropriately labelled tube and allow it to clot. Centrifuge and carefully remove the serum layer. Store at 4°C for up to 24 hours or at -10°C or lower if the analyses are to be done at a later date. 
Plasma: Approximately 0.2 mL of plasma is required per duplicate determination. Collect 4–5 mL of blood into EDTA plasma tubes. Store at 4°C for up to 24 hours or at -10°C or lower if the analyses are to be done at a later date. 
Urine: 24-hour urine into a specimen collection container. Store at 4°C for up to 24 hours or at -10°C or lower if the analyses are to be done at a later date. 
Consider all human specimens as possible biohazardous materials and take appropriate precautions when handling.

CALCULATIONS 
1. Calculate the mean optical density of each calibrator duplicate. 
2. Draw a calibrator curve on semi-log paper with the mean optical densities on the Y-axis and the calibrator concentrations on the X-axis. If immunoassay software is being used, a 4-parameter or 5-parameter curve is recommended. 
3. Calculate the mean optical density of each unknown duplicate. 
4. Read the values of the serum and plasma samples directly off the calibrator curve. 
5. Read the values of the urine samples directly off the curve and multiply by a factor of 50. Next, multiply by the volume of collected 24-hour urine (in mL) to obtain values in pg/24 hour. Finally, divide the pg/24 hour values by 1x106 (1,000,000) to obtain values in μg/24 hour. 
6. If a serum or plasma sample reads greater than 1000 pg/mL then dilute it with the Serum and Plasma Diluent (available separately under REF CAN-ALD-500-11) at a dilution of no more than 1:8. The result obtained must be multiplied by the dilution factor. If a urine sample reads more than 1000 pg/mL then dilute it with the urine diluent at a dilution of no more than 1:8 (from the original 1:50 dilution). The result obtained must be multiplied by the dilution factor.