Hypercalcemia and Hypocalcemia - Vitamin D Testing

Medical Experts

Contributor

Doyle

Kelly Doyle, PhD, DABCC, FAACC

Associate Professor of Pathology (Clinical), University of Utah

Medical Director, Special Chemistry and Endocrinology, ARUP Laboratories

Contributor

Rudolf

Joseph Rudolf, MD

Assistant Professor of Pathology (Clinical), University of Utah

Medical Director, Automated Core Lab, ARUP Laboratories

Contributor

Straseski

Joely A. Straseski, PhD, MS, MT(ASCP), DABCC, FAACC

Professor of Pathology (Clinical), University of Utah

Head of Clinical Operations for Clinical Chemistry, Toxicology, and Biochemical Genetics; Medical Director, Endocrinology; Co-Director, Automated Core Laboratory, ARUP Laboratories

Calcium imbalance (hyper- or hypocalcemia) is a metabolic abnormality with widespread effects that may be acute or chronic. Mild cases may be asymptomatic, whereas acute, severe hyper- or hypocalcemia may present with musculoskeletal, gastrointestinal, cardiovascular, neurologic, or psychiatric symptoms. Serum calcium exists as free (ionized), protein bound, or chelated. The major binding proteins include albumin and globulin. Low albumin can affect serum calcium; thus, accurate serum calcium testing requires correction with a concurrent albumin concentration. Ionized calcium, which comprises about half of the total serum calcium, is the form used to maintain physiologic functions. Testing for ionized calcium can remove variability due to albumin or confirm a possible abnormal value. Given the complex regulation of calcium homeostasis in the body, when evaluating for hyper- or hypocalcemia, laboratory testing for albumin, phosphorous, magnesium, creatinine, parathyroid hormone (PTH), and vitamin D (25-hydroxyvitamin D) should also be performed.

Quick Answers for Clinicians

What is the correct vitamin D test to order in the investigation of vitamin D deficiency?

The recommended test to evaluate for vitamin D deficiency is the 25-hydroxyvitamin D [25(OH)D] test, as it most accurately reflects the concentration of vitamin D. Vitamin D is composed of vitamin D₂, ergocalciferol, and vitamin D₃, cholecalciferol. Individual testing for vitamin D₂ and D₃ may be useful as follow-up testing in individuals who do not respond to therapeutic vitamin D supplementation. Conversion of 25(OH)D to the active metabolite, 1,25-dihydroxyvitamin D [1,25(OH)₂D], via 1 alpha (α)-hydroxylase, occurs mainly in the kidneys. As only a fraction of 25(OH)D is converted to 1,25(OH)₂D, measurement of 1,25(OH)₂D does not reflect accurate vitamin D concentrations and is only useful in cases of inherited or acquired dysregulation of phosphorous and vitamin D metabolism, such as chronic kidney disease, oncogenic osteomalacia, and vitamin D-resistant rickets.

What are the appropriate reference intervals for vitamin D tests?

The U.S. Preventive Services Task Force (USPSTF) and the Endocrine Society do not endorse any specific reference intervals for vitamin D deficiency, insufficiency, or sufficiency.^, The Endocrine Society previously recommended defining sufficiency as 30 ng/mL, insufficiency as 21-29 ng/mL, and deficiency as <20 ng/mL; however, there was a lack of evidence from clinical trials to support these values.^, Individuals have varying biologic requirements for vitamin D. Low vitamin D concentrations have been shown to increase the risk for autoimmune, metabolic, cardiovascular, malignant, infectious, and musculoskeletal diseases; however, causative relationships have not been established.

What testing is recommended for suspected hypercalcemia of malignancy?

Malignancy is a common cause of hypercalcemia, particularly in those with multiple myeloma or solid tumors such as lung, breast, and renal cancers. Hypercalcemia in these cases is due to the production of parathyroid hormone-related peptide (PTHrP), 1,25(OH)D₂, or metastases to the bone inducing the release of osteoclast-activating factors. In cases of confirmed hypercalcemia in which the PTH is suppressed and symptoms have a rapid onset, workup for malignancy should be considered, including testing for PTHrP, 1,25(OH)D₂, and alkaline phosphatase. Testing for PTHrP may be indicated in other conditions that do not involve malignancy, such as pregnancy, systemic lupus erythematosus, benign ovarian and kidney tumors, and HIV-associated lymphadenopathy.

What is the role of ionized calcium testing in hypocalcemia and hypercalcemia?

Because a portion of total serum calcium is bound to albumin, low albumin concentrations may alter measured total serum calcium results. Testing for ionized calcium, the physiologically active form, can remove the variability caused by albumin to confirm an abnormal value. Ionized calcium does not need to be corrected for albumin, although it does need to be corrected for pH (corrected ionized calcium = measured iCa²⁺ [1–0.53 × (7.40–measured pH)]). Measurement of ionized calcium is more accurate in patients with chronic kidney disease, hyperparathyroidism, neonatal hypocalcemia, hypercalcemia of malignancy, and critically ill patients undergoing surgery.

Indications for Testing

Laboratory testing for calcium and vitamin D is used to:

Investigate signs and symptoms of hypocalcemia (eg, arrhythmias, perioral numbness, paresthesias, muscle spasms/cramps, Chvostek sign, Trousseau sign)
Investigate signs and symptoms of hypercalcemia (eg, arrhythmias, constipation, arthralgias, myalgias, bone pain, lethargy, mental status changes, anxiety, recurrent nephrolithiasis)^,
Follow up on calcium concentration after thyroidectomy or parathyroidectomy^,
Assess calcium homeostasis in calcium dysregulation disorders (eg, chronic kidney disease [CKD], primary hyperparathyroidism, hypoparathyroidism)
Investigate signs and symptoms of vitamin D deficiency (eg, hypocalcemia, rickets, osteomalacia, osteoporosis, nontraumatic fractures)
Screen for vitamin D deficiency in individuals with specific risk factors (refer to the Screening section for more information)
Monitor therapy (both treatment and medications that increase the risk of hypo- or hypercalcemia and vitamin D deficiency)

Laboratory Testing

Screening

The U.S. Preventive Services Task Force (USPSTF) finds insufficient evidence to make a recommendation for or against vitamin D screening in asymptomatic, community-dwelling, nonpregnant adults. The Endocrine Society recommends against routine 25(OH)D testing in individuals without an established indication, as no clinical trials have shown outcome-specific benefits.

Screening should be performed in individuals at risk, such as those with CKD, nephrotic syndrome, hepatic failure, malabsorption syndromes, or a history of bariatric surgery.^,^, The USPSTF also includes older age and obesity as risk factors for vitamin D deficiency that warrant screening. The Endocrine Society does not endorse screening in older individuals or those with obesity due to a lack of clinical data.

Diagnosis

Initial Evaluation

Initial testing for calcium imbalance involves measurement of calcium, albumin, phosphorous, magnesium, creatinine, PTH, and vitamin D. Albumin, phosphorous, magnesium, and creatinine concentrations may inform the interpretation of calcium test results and the etiology of the imbalance.

A portion of serum calcium is bound to albumin, which causes serum calcium concentrations to be affected by varying albumin concentrations. To account for the effects of albumin, the following formula may be used to calculate a corrected calcium value: corrected calcium (mg/dL) = serum calcium (mg/dL) + 0.8 (4.0 g/dL - serum albumin g/dL).

If the corrected calcium is low or borderline, a repeat test with ionized calcium may be useful because ionized calcium does not need to be corrected for albumin (although it does need to be corrected for pH).

Parathyroid Hormone Testing

If hypo- or hypercalcemia is confirmed, and/or if the patient is symptomatic, intact PTH should be tested. In addition to PTH testing, phosphorous, magnesium, creatinine, vitamin D, and 24-hour urinary calcium and creatinine should be measured to aid in the diagnostic workup. Refer to the testing algorithms for hypocalcemia and hypercalcemia for more information.

Interpretation of Calcium and PTH Test Results
Calcium^a	Intact PTH	Potential Interpretations
Low	Low/normal	Hypoparathyroidism Magnesium deficiency
Low	High	Pseudohypoparathyroidism Vitamin D deficiency Chronic kidney disease-mineral bone disorder Secondary hyperparathyroidism (eg, CKD)
High	Low	Malignancy^b Lymphoma Granulomatous disease Vitamin D excess Milk-alkali syndrome
High	Normal or high	Primary hyperparathyroidism Familial hypocalciuric hypercalcemia
^aReference intervals depend on the particular test used and the age of the patient. ^bApproximately 90% of cases of hypercalcemia are linked to primary hyperparathyroidism and malignancy. Sources: AAFP, 2003; ASMBR, 2022

Vitamin D Testing

25-Hydroxyvitamin D

Testing for total 25-hydroxyvitamin D [25(OH)D] is the preferred test to screen for, diagnose, and monitor vitamin D deficiency in those who have an indication.^, Testing for 25(OH)D can be used to identify vitamin D deficiency as the etiology of hypocalcemia or hyperparathyroidism.^, The concentration of 25(OH)D in the blood reflects vitamin D stores in the liver, and 25(OH)D is stable.

D₂(ergocalciferol) and D₃(cholecalciferol) make up 25(OH)D, and both D₂ and D₃ are measured by total 25(OH)D tests. D₂ and D₃ concentrations can be measured separately. Separate testing of D₂ and D₃ is not necessary in the evaluation of vitamin D deficiency but may be useful in individuals who are not responsive to therapeutic supplementation.

1,25-Dihydroxyvitamin D

Although 1,25 dihydroxyvitamin D [1,25(OH)₂D] is the bioactive form of vitamin D, it is unstable due to its short half-life. Only a fraction of 25(OH)D is converted to 1,25(OH)₂D, which means 1,25(OH)₂D concentrations do not accurately reflect vitamin D stores.^, Measurement of 1,25(OH)₂D may be useful in cases of inherited or acquired dysregulation of phosphorous and vitamin D metabolism.

Other Tests

A renal function panel may be useful for evaluating calcium metabolism in individuals with hypercalcemia due to secondary hyperparathyroidism, in addition to 25(OH)D, calcium, phosphorous, and PTH testing.

Measurement of 24-hour urinary calcium and creatinine may be helpful in the workup of hypercalcemia, as the ratio may be used to distinguish benign familial hypercalcemia/familial hypocalciuric hypocalcemia from primary hyperparathyroidism. Measurement of urinary calcium is also indicated annually in those with a history of renal calculi in the setting of asymptomatic primary hyperparathyroidism. In patients with nephrolithiasis, it may be useful to measure urinary calcium, as hypercalciuria is a significant cause of idiopathic calcium-based stones.

Monitoring

Monitoring of hypocalcemia treatment should include weekly to monthly measurements of serum calcium, phosphorous, and creatinine during initial therapy. Once values have stabilized during therapy, monitoring becomes less frequent. Serum calcium, phosphorous, magnesium, and creatinine should be measured every 3-6 months. Serum 25(OH)D, urine calcium, creatinine, and sodium should be measured every 6-12 months.