Calcium Examination Procedure

1.Purpose of examination: Calcium estimation from serum or plasma by Arsenazo III Method.

2.Responsibility and Authority:

• Calibration: Technician
• Quality Control: Technician
• Routine operation: Technician
• Overall Monitoring: Quality Manager

3.Sample Details:

• Type of Sample: Serum, Plasma
• Type of container and additives: Plain without any additives
• Patient Preparation: As per Primary Sample Collection Manual sample_collection_manual
• Stability: At Room temperature 18°–28°C (64°–82°F) stability ≤ 24 hours
• Handling and transport: As per Primary Sample collection manual
• Storage: 24 hours at 2-8° C

4.Required Equipment: Centrifuge, Auto-Pipette, Disposable Tips, Disposable sample cups, FullyAuto Chemistry Analyzer

5.Required reagents: R1 Arsenazo-III dye 0.94 mmol/L Sodium acetate 271 mmol/L

6.Reagent Handling Remove any air bubbles present in the reagents with a new applicator stick, or allow the reagents to settle at the appropriate storage temperature to allow the bubbles to dissipate. To minimize volume depletion, do not use a transfer pipette to remove bubbles

7.Reagent Storage and stability

• Unopened reagent stable at 2-8°C until expiration date.
• On board System temperature reagent is stable for 30 days
• Instability or deterioration should be suspected if there are precipitates, visible signs of leakage or
• contamination, turbidity, or if calibration or controls do not meet the appropriate criteria.

8.Calibration Procedure:

• Multiconstituent Consolidated Calibrator(MCC)
• Frequency:

1. Reagent lot change
2. QC out of range
3. After service or maintenance
4. Replacement in any parts of Instrument

Procedure:

1. Start the equipment.WDI abbotte fully.docx
2. Calibrators are ready to use.
3. Put calibrator 15-20 minutes at room temperature
4. Prior to use, mix the contents by inverting the vial. Do not shake the vial to prevent foam formation.
5. Take a 150 µl both level of calibrator solution in to separate aliquot.
6. Go to the calibration and give the calibration order.
7. Verify calibration with at least two levels of controls
8. If control results fall outside acceptable ranges,root cause analysis or recalibration may be necessary.

9.Quality control Procedure: Name: Biorad Level 1 & 2 Frequency: As per Quality Control Procedure

Procedure for Reconstitution of IQC

1. Reconstitution of QC material with 5 ml Distilled water by using calibrated fixed volume pipette.
2. Leave to stand for 30 min in the dark place.
3. Swirl gently several times during the reconstitution period to ensure that the contents are completely dissolved.
4. Prior to use, mix the contents by inverting the vial. Do not shake the vial as the information of foam should be avoided. Ensure that no lyophilized material remains un-reconstituted.
5. Prepare aliquots of 150 µl from the reconstituted QC material.
6. Store these aliquots at -15° C to -20° C.
7. Prior to use, make sure that aliquots should be at room temperature for at least 15 min.

Procedure to run IQC

• Press Control order
• Select Assay /Panel, to be run.
• Select the control/s and its level/s
• Give Carrier Number and Position number
• Press F3 / Add order
• Put respected carrier in RSH rack
• Check IQC results, in case outliers call residents.

10.Principle of the procedure used for examinations: Arsenazo-III dye reacts with calcium in an acid solution to form a blue-purple complex. The color developed is measured at 660 nm and is proportional to the calcium concentration in the sample.

11.Sample Preparation:

1. Required SampleVolume: 150 µl of the sample
2. Temperature: 37°

Take 150-200µl of the sample from Primary tube to the secondary aliquot. Write the sample ID on the aliquot.

Procedure to run Patient sample

• Press Patient order
• Select Assay /Panel, to be run.
• Give Carrier Number and Position number
• Press F3 / Add order
• Put respected carrier in RSH rack

12.Performance Characteristics:

• Linearity: 2.0 to 24.0 mg/dL
• Limit of Detection (LOD): 0.5 mg/dL
• Limit of Quantitation (LOQ):1.0 mg/dL
• Unit: mg/dl

Automated Dilution Protocol: When using the Automated Dilution Protocol, the system performs a dilution of the specimen and automatically corrects the concentration by multiplying the result by the appropriate dilution factor.

Manual Dilution Procedure: Dilute the specimen with saline (0.85% to 0.90% NaCl). Enter the dilution factor in the Patient or Control order screen.

Normal and critical ranges:

Calcium, total

13.Laboratory Clinical interpretation:

• Hypercalcemia can result from hyperparathyroidism, hypervitaminosis D, multiple myeloma, and some neoplastic diseases of bone.1 Long- term lithium therapy has been reported to cause hyperparathyroidism in some individuals, with resulting hypercalcemia.
• Hypocalcemia can result from hypoparathyroidism, hypoalbuminemia, renal insufficiency, and pancreatitis.1

14.Interference and cross reaction:

• Bilirubin 30 mg/dL (513 μmol/L)
• Hemoglobin 1,000 mg/dL
• Intralipid 500 mg/dL

15.Potential source of variation: Turn around time (TAT):

• Routine: 6.0 hours
• Urgent: 2.0 hours

16.Recording of observation:

• Software backup
• Machine raw data

17.Storage & Disposal of waste: Follow storage & discard procedure

18.Environmental & Safety control: Reagent R1 contains sufamic acid that Causes severe skin burns and eye Damage

19.Precautions:

• Avoid breathing mist / vapors / spray.
• Wash hands thoroughly after handling.
• Contaminated work clothing should not be allowed out of the workplace.
• Wear protective gloves / protective clothing / eye protection.

20.Response

1. IF IN EYES: Rinse cautiously with water for several minutes. Remove contact lenses, if present and easy to do. Continue rinsing.
2. If eye irritation persists: Get medical advice / attention.
3. IF ON SKIN: Wash with plenty of water.If skin irritation or rash occurs: Get medical advice / attention. Take off contaminated clothing and wash it before reuse.

21.References:

1. Bendz H, Sjödin I, Toss G, et al. Hyperparathyroidism and long-term lithium therapy—a cross-sectional study and the effect of lithium withdrawal. J Intern Med 1996;240:357–365.
2. US Department of Labor, Occupational Safety and Health Administration, 29 CFR Part 1910.1030, Bloodborne pathogens.
3. US Department of Health and Human Services. Biosafety in Microbiological and Biomedical Laboratories. 5th ed. Washington, DC: US Government Printing Office; December 2009.
4. World Health Organization. Laboratory Biosafety Manual. 3rd ed. Geneva: World Health Organization; 2004.