STANDARD OPERATING PROCEDURE (SOP) FOR ESTIMATION OF DIETARY INTAKE OF SODIUM (DIS) IN 24 HOUR URINE USING ELECTROLYTE ANALYZER

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1 STANDARD OPERATING PROCEDURE (SOP) FOR ESTIMATION OF DIETARY INTAKE OF SODIUM (DIS) IN 24 HOUR URINE USING ELECTROLYTE ANALYZER Estimation of Dietary Intake of Sodium (DIS) in 24h Urine on electrolyte analyzer A. Principle: The electrolyte analyzer has different electrodes, specific for different ions of interest. Each electrode has an ion selective membrane (for example for sodium) that undergoes a specific reaction with the corresponding ions (sodium) contained in the sample being analyzed. The membrane is an ion exchanger reacting to the electric charge of the ion causing a change in the membrane potential or the measuring voltage, which is built up in the film between the sample and the membrane. A difference in sodium ion concentration between the sodium solution inside the electrode and the sample causes an electrochemical potential to form across the membrane of the active electrode. The potential is conducted by the electrode to an amplifier. This is compared with the potential of a reference electrode. The sodium ion concentration measured is expressed in terms of mmol/l or meq/l. B. Equipment and Materials required: 1. Electrolyte analyzer automated microprocessor controlled analytic instrument based on Ion Selective Electrode (ISE) technology 2. Reagent pack for calibration 3. Waste storage chamber for collecting waste 4. Urine Diluent 5. Na Conditioner 6. Quality Control (QC) solution provided by the manufacturer of electrolyte analyzer. 1

2 7. Urine based internal QC solution: The controls come in the lyophilized form (freezedried) for enhanced stability. 100% human urine based control should be run. Reconstituted stability after reconstituting urine control is generally of 5 days at 2-8ºC or 14 days at -20ºC. 8. Daily cleaning solution 9. Weekly cleaning solution: 10. Auto pipette µl with pipette tips 11. 2ml eppendorf tube for sample dilution C. Methodology The methodology will vary with different electrolyte analyzer. The method described here is for performing sodium analysis in Electrolyte Analyzer from Caertium Step 1: Sample Preparation: 1. Urine sample, if stored at 4 o C or -20 o C should be allowed to come to room temperature. 2. Urine aliquot should be mixed properly to ensure homogeneity. 3. Aspirate 250 µl of urine into a 2 ml tube using micropipette and to this add 250µl of urine diluent. 4. Mix properly by inverting 5-10 times. STEP 2: Installation of the reagent pack and start up of the instrument. 1. Position the reagent pack into the groove of the instrument 2. Turn on the power and boot up the instrument 3. The instrument carries out self-test. 4. After this instrument carries out self-calibration at the end of which the slope of the electrodes will be displayed on the screen and also printed out. Check to see if the slope 2

3 is in normal range (Should be between mv for sodium). OK is displayed if the values are between mv. Otherwise recalibration, after cleaning the electrode would be required. 5. After the calibration the main menu is displayed. Step 3: Running the quality check Quality check should be done every day 1. Press the tab service in the main menu to display the service menu and then press the tab QC to enter into measurement menu. 2. Present the quality control solution provided with the instrument under the sample probe and press the tab aspirate, the result will be displayed and printed in 60 seconds. 3. This value should be within the range specified on the vial of the quality control. For example for sodium the value given on the vial may be 145±2 mmol/l. So if the value of QC is between mmol/l we will move on to the next step. 4. When more than 5 QC results are available, a statistical report can be obtained to see the trend. 5. Press the tab stat on the QC measurement menu to get mean and standard deviation of QC values. Step 4: Preparation of work list for running urine samples 1. A work list should be prepared if more than one sample is being run. This is for convenience. 2. Press the tab W. list to enter into input number program. Press the tab U/L to change the mode of measurement to urine. The operator can enter the sample number and the patient ID manually to create a programme for the number of samples to be analyzed. Step 5: Testing of urine control 1. Press the tab sample to enter into the sample measurement menu. 3

4 2. Present the diluted urine control (prepared as mentioned in step 1) under the sample probe and press the tab aspirate, the sample will be aspirated into the system. 3. Remove the sample when the screen shows remove sample with a beep sound. 4. The result will be displayed within 55 seconds and printed out. Repeat the process to measure the remaining samples. 5. If the urine quality control value is within the range specified for the control we can proceed to the next step. Step 6: Testing of urine sample 1. Press the tab sample to enter into the sample measurement menu. 2. Present the diluted urine sample (prepared as mentioned in step 1) under the sample probe and press the tab aspirate, the sample will be aspirated into the system. 3. Remove the sample when the screen shows remove sample with a beep sound. 4. The result will be displayed within 55 seconds and printed out. Repeat the process to measure the remaining samples. Calculation of DIS in 24 hrs: For each individual, the 24 h sodium excretion value (mmol/day) is calculated as the concentration of sodium in the urine (mmol/l) multiplied by the total urinary volume (L/day). D. Quality assurance External Quality Assessment (EQA): External quality assessment (EQA) permits comparison of results between laboratories measuring the urinary sodium. EQA is a retrospective process of assessment of performance, particularly of inaccuracy or bias with respect to mean values. EQC program for sodium is available through different sources. Example: 1. Biorad Urine chemistry Program. 4

5 2. Wales External Quality Assessment Scheme (WEQAS) Both EQC programs send 12 samples/year. E. Maintenance of the electrolyte analyzer: Daily maintenance: 1. Check the reagent residual volume and replace as and when needed. 2. At the end of analysis for the day, run the daily maintenance solution. Weekly maintenance: 1. Check the voltage of each electrode. Replace the reference filling solution in electrode or the reference membrane, if necessary. Refill the filling solution in the electrode if the volume is less than 2/3 of the total volume. 2. If there is salty crystal on the electrode, clean it with wet tampon. 3. Run cleaning program in service menu once a week if more than 25 samples measured every day. If less than 20 samples measured every day, then the investigator just need to run the cleaning program every 2-3 weeks. 4. Run Na adjust program if the slope of Na electrode is less than 45. For further problems the service engineer should be contacted. 5