What tests should be ordered next for suspected vitamin B12 deficiency?

Testing Strategy for Suspected Vitamin B12 Deficiency

Start with total serum vitamin B12 as your first-line test, then add methylmalonic acid (MMA) only when B12 results fall in the indeterminate range (180-350 pg/mL) to confirm functional deficiency. 1

Initial Laboratory Assessment

Order total serum vitamin B12 first because it costs approximately £2 per test with rapid turnaround in local laboratories, making it the most cost-effective initial screening tool. 1 Active B12 (holotranscobalamin) is more accurate at measuring the biologically active form available for cellular use, but costs £18 per test with longer processing times, making it less practical for routine initial screening. 1

Interpretation of Total Serum B12

• < 180 pg/mL (< 133 pmol/L): Confirms deficiency—initiate treatment immediately without additional testing. 1, 2
• 180-350 pg/mL (133-258 pmol/L): Indeterminate range—proceed to MMA measurement to identify functional deficiency. 1, 2
• > 350 pg/mL (> 258 pmol/L): Deficiency unlikely, but consider MMA testing if clinical suspicion remains high (neurological symptoms, high-risk conditions). 1

When to Order Methylmalonic Acid (MMA)

MMA should be measured only when initial B12 results are indeterminate (180-350 pg/mL), not as a first-line screening test. 1 This targeted approach is cost-effective at £3,946 per quality-adjusted life year, whereas universal MMA screening wastes resources due to the £11-80 per test cost and specialized equipment requirements. 1

Why MMA Matters

MMA detects an additional 5-10% of patients with functional B12 deficiency who have low-normal serum B12 levels. 1 In the Framingham Study, 12% had low serum B12, but an additional 50% had elevated MMA indicating metabolic deficiency despite "normal" serum levels. 1 In polyneuropathy patients, 44% had B12 deficiency based solely on abnormal metabolites when serum B12 was normal. 1

MMA Interpretation

• MMA > 271 nmol/L: Confirms functional B12 deficiency with 98.4% sensitivity. 1
• MMA is more specific than homocysteine for B12 deficiency—MMA is elevated in only 12.2% of folate-deficient patients versus 91% for homocysteine. 1

Critical Caveat for MMA

Both MMA and homocysteine can be falsely elevated in hypothyroidism, renal insufficiency, and hypovolemia—interpret cautiously in these conditions. 1

Additional Testing to Consider

Complete Blood Count (CBC)

Order a CBC to check for megaloblastic anemia, though this is absent in one-third of B12 deficiency cases. 1 Macrocytosis (elevated MCV) often appears before anemia develops, making it an early laboratory sign. 1

Homocysteine (Optional)

Homocysteine > 15 μmol/L supports B12 deficiency diagnosis, though it is less specific than MMA and can be affected by renal impairment and other B vitamins. 1 The pattern helps differentiate causes:

• Elevated homocysteine + elevated MMA = B12 deficiency 1
• Elevated homocysteine + normal MMA = folate deficiency 1

High-Risk Populations Requiring Testing

Even if you haven't ordered B12 yet, consider testing in patients with these risk factors:

• Age > 75 years: 18.1% have metabolic deficiency; 25% of those ≥ 85 years have B12 < 170 pmol/L. 1
• Metformin use > 4 months 1, 3
• PPI or H2 blocker use > 12 months 1
• Gastric or ileal resection (especially > 20 cm) 1, 3
• Inflammatory bowel disease (Crohn's with ileal involvement) 1, 3
• Vegans or strict vegetarians 1, 3
• Post-bariatric surgery patients 1
• Autoimmune thyroid disease: 28-68% prevalence of B12 deficiency. 1

Alternative: Active B12 (Holotranscobalamin) as Initial Test

If you choose active B12 instead of total B12:

• < 25 pmol/L: Confirms deficiency—treat immediately. 1
• 25-70 pmol/L: Indeterminate—order MMA. 1
• > 70 pmol/L: Deficiency unlikely. 1

Active B12 is more sensitive and measures only the biologically active form, but the higher cost and longer turnaround time make total serum B12 the preferred initial test in most clinical settings. 1

What NOT to Do

Do not order MMA as a first-line screening test because it is not cost-effective due to expense and processing delays. 1 Use it only to clarify indeterminate B12 results.

Do not rely solely on serum B12 to rule out deficiency in patients > 60 years, where metabolic deficiency is common despite normal serum levels. 1 Standard serum B12 testing misses functional deficiency in up to 50% of cases. 1

Do not stop at one normal B12 result if clinical suspicion remains high—consider MMA testing, especially in elderly patients or those with neurological symptoms. 1

Practical Algorithm Summary

1. Order total serum vitamin B12 first (costs £2, rapid results). 1
2. If B12 < 180 pg/mL: Diagnose deficiency and treat immediately. 1
3. If B12 180-350 pg/mL: Order MMA; if MMA > 271 nmol/L, confirm functional deficiency and treat. 1
4. If B12 > 350 pg/mL but high clinical suspicion: Consider MMA on a case-by-case basis. 1
5. Add CBC to check for macrocytosis or megaloblastic anemia. 1
6. Consider homocysteine if you need to differentiate B12 from folate deficiency. 1