Do I need to refrigerate the sample during a 24-hour urine calcium test?

24-Hour Urine Calcium Collection: Refrigeration Requirements

Yes, you must refrigerate your 24-hour urine collection sample throughout the entire collection period to ensure accurate results and prevent bacterial overgrowth. 1, 2

Storage Protocol During Collection

Keep the collection container refrigerated at 4°C (approximately 39°F) continuously during the entire 24-hour collection period. 1, 3 This is the standard recommendation for maintaining specimen integrity when processing cannot occur immediately.

Why Refrigeration Matters

• Bacterial overgrowth prevention: Urine kept at room temperature for more than 1 hour allows bacterial proliferation, which can alter calcium measurements and other analytes. 1, 2

• Specimen stability: Refrigeration at 4-10°C maintains the stability of calcium and other urinary constituents for up to 24 hours without significant degradation. 3, 4

• Diagnostic accuracy: Unrefrigerated specimens beyond 1 hour may produce false results, compromising the clinical interpretation of your calcium excretion status. 1, 2

Practical Collection Instructions

Follow this algorithm for proper 24-hour urine collection:

1. Start collection: Discard the first morning void, note the time. 1

2. During collection: Store the collection container in the refrigerator between voids. 1, 3

3. Each void: Remove container from refrigerator, add urine, immediately return to refrigerator. 1

4. Complete collection: Include the first morning void 24 hours after starting, then transport to laboratory promptly. 1, 3

Important Considerations About Acidification

You do NOT need to add acid to your urine collection for calcium measurement. 5, 6 This is a critical point that contradicts older practices:

• Recent high-quality research demonstrates that acidification (either before or after collection) does not significantly affect calcium measurement accuracy. 5, 6

• In a study of 567 urine samples, only 4.4% showed analytical differences with acidification, and these differences did not clinically reclassify patients. 5

• Another study of 133 patients found no significant difference between acidified and non-acidified samples for calcium measurement. 6

• The most recent 2022 evidence confirms that preservation with acid is unnecessary for storage up to 72 hours when samples are properly refrigerated. 4

Common Pitfalls to Avoid

• Never leave the collection container at room temperature for extended periods between voids, as this allows bacterial growth that can interfere with multiple analytes. 1, 2

• Do not freeze the sample during collection, as this can damage cellular elements and alter measurements. 1

• Do not add acid unless specifically instructed by your laboratory, as modern calcium assays do not require acidification and acid poses unnecessary safety risks. 5, 6

• Transport promptly after completion: Once the 24-hour collection is complete, deliver to the laboratory as soon as possible, keeping refrigerated during transport if there will be any delay. 1, 3

Temperature-Specific Evidence

The evidence strongly supports refrigeration over room temperature storage:

• Room temperature storage beyond 1 hour causes bacterial overgrowth and diagnostic errors in up to 32% of specimens. 2

• Refrigerated storage at 4°C maintains specimen integrity for 24-72 hours for calcium and other stone risk factors without significant degradation. 4, 7

• Studies comparing different storage conditions found no significant deviation in calcium, magnesium, phosphorus, oxalate, citrate, or uric acid when samples were refrigerated for up to 72 hours. 4