What laboratory tests are recommended for a patient suspected of having a mast cell disorder?

Laboratory Testing for Suspected Mast Cell Disorders

Start with serum tryptase measurement and peripheral blood KIT D816V mutation testing using highly sensitive ASO-qPCR, followed by bone marrow biopsy if tryptase is persistently >20 ng/mL or if systemic mastocytosis is suspected. 1, 2

Initial Laboratory Workup

For all patients with suspected mast cell disorders, obtain the following baseline tests:

• Serum tryptase (baseline) when the patient is completely asymptomatic to establish their personal reference value 2, 3
• Peripheral blood KIT D816V mutation using highly sensitive allele-specific oligonucleotide quantitative PCR (ASO-qPCR) if baseline tryptase >20 ng/mL 1, 2
• Complete blood count with differential to assess for eosinophilia or other hematologic abnormalities 2
• 24-hour urine collection for N-methylhistamine, leukotriene E4 (LTE4), and 11β-prostaglandin F2α (11β-PGF2α) as complementary markers 2, 3

Critical caveat: Do NOT use plasma or urine histamine levels, as these are not validated markers; N-methylhistamine is the preferred histamine metabolite 3. Similarly, heparin and chromogranin A are not reliable markers for mast cell activation 3.

Testing During Symptomatic Episodes (For MCAS Diagnosis)

If mast cell activation syndrome is suspected, timing of specimen collection is crucial:

• Acute serum tryptase drawn 1-4 hours after symptom onset during an active episode 2, 3, 4
• Diagnostic threshold: ≥20% increase above the patient's baseline tryptase PLUS an absolute increase of ≥2 ng/mL 2, 3, 4
• Spot urine collection during symptomatic episodes for mediator metabolites if serum tryptase is difficult to obtain 2, 3

The paired tryptase measurement (baseline vs. acute) is essential because comparing acute to baseline levels optimizes detection of mast cell activation 3. A single elevated tryptase without comparison to baseline is insufficient for MCAS diagnosis.

Bone Marrow Evaluation

Proceed to bone marrow biopsy and aspirate when:

• Baseline serum tryptase persistently >20 ng/mL 1, 3
• Peripheral blood KIT D816V mutation is positive 1
• Clinical features suggest systemic mastocytosis (organomegaly, abnormal blood counts, adult-onset mastocytosis in skin) 2, 3
• Suboptimal bone marrow aspirate (dry tap) requires alternative peripheral blood testing 1

Bone marrow analysis must include:

• Core biopsy to assess for multifocal dense mast cell infiltrates (major diagnostic criterion) 1
• Immunohistochemistry with mast cell tryptase, CD117, and CD25 to quantify mast cell burden 1
• Flow cytometry for aberrant CD25 and CD2 expression on mast cells (standard markers); CD30 is optional 1, 2
• KIT D816V mutation analysis on bone marrow aspirate if peripheral blood is negative 1
• Reticulin and collagen staining to assess bone marrow fibrosis grade (MF-0 to MF-3) 1
• Chromosome analysis especially if associated hematologic neoplasm is suspected 1
• Myeloid mutation panel on bone marrow (or peripheral blood if AHN present) 1

Important technical consideration: KIT mutational analysis can be performed on formalin-fixed paraffin-embedded tissue only if it has not been decalcified or has been decalcified in EDTA; other fixatives and rapid decalcification yield unsatisfactory results 1.

Special Genetic Testing

For patients with interstitial mast cell pattern, peripheral eosinophilia, and negative KIT D816V:

• Test for FIP1L1-PDGFRA fusion gene to exclude hypereosinophilic syndrome variants 1

For hereditary considerations:

• Buccal swab for TPSAB1 α-tryptase copy number variation (CNV) to diagnose hereditary α-tryptasemia, which can elevate baseline tryptase and confound interpretation 3

If KIT D816V remains negative despite high mast cell burden:

• Confirm with the most sensitive ASO-qPCR technique available 1
• Evaluate for alternative codon 816 mutations (D816H, D816Y, others) or mutations in other KIT regions 1
• Consider testing skin or extracutaneous organs besides bone marrow in low-burden cases 1

Sensitivity and Limitations

Understanding test performance prevents misdiagnosis:

• ASO-qPCR detects KIT D816V in >80% of systemic mastocytosis patients when applied to bone marrow 1
• In 5-10% of patients, no KIT D816V is detected due to: (1) very low mast cell burden causing false-negative results, (2) true wild-type KIT, or (3) presence of other KIT mutations not detected by standard ASO-qPCR 1
• Next-generation sequencing (NGS) myeloid panels have only ~5% sensitivity for KIT D816V and are NOT recommended for this purpose 1

Practical Algorithm Summary

Step 1: Baseline serum tryptase + peripheral blood KIT D816V (if tryptase >20 ng/mL) + 24-hour urine mediators 2, 3

Step 2: If symptomatic episodes occur, obtain acute serum tryptase 1-4 hours after onset and compare to baseline 2, 3, 4

Step 3: If tryptase persistently >20 ng/mL or KIT D816V positive, proceed to bone marrow biopsy with comprehensive analysis 1, 2

Step 4: If bone marrow shows primarily interstitial pattern with eosinophilia and negative KIT D816V, test for FIP1L1-PDGFRA 1