Laboratory Tests for HLH Diagnosis
The diagnosis of HLH requires either molecular/genetic testing confirming HLH-associated mutations OR fulfillment of 5 out of 8 specific laboratory and clinical criteria from the HLH-2004 protocol. 1, 2, 3
Core Laboratory Panel (HLH-2004 Criteria)
The following laboratory tests must be obtained when HLH is suspected, with 5 of 8 criteria required for diagnosis:
Essential Laboratory Tests
Complete blood count with differential: Look for cytopenias affecting ≥2 of 3 lineages (hemoglobin <9 g/dL, platelets <100 × 10⁹/L, neutrophils <1.0 × 10⁹/L) 4, 1, 3
Ferritin: Must be ≥500 ng/mL per HLH-2004 criteria, though values >7,000-10,000 ng/mL (sometimes >100,000 ng/mL) are more characteristic of HLH in adults 4, 3
Triglycerides: Hypertriglyceridemia defined as ≥265 mg/dL (≥3.0 mmol/L) 1, 3
Fibrinogen: Hypofibrinogenemia defined as ≤150 mg/dL (≤1.5 g/L) 1, 3
Soluble IL-2 receptor (sCD25): Must be ≥2,400 U/mL; this test has excellent diagnostic performance with an area under the curve of 0.90 compared to 0.78 for ferritin 4, 3
NK cell activity: Low or absent NK cell activity is a key immunological criterion 1, 2
Tissue-Based Testing
- Bone marrow aspirate and biopsy: Examine for hemophagocytosis, though this finding is neither sensitive nor specific and may be absent initially 4, 3
Additional Supportive Laboratory Tests
Liver function tests: Check AST, ALT, bilirubin, and albumin for hepatic involvement 4, 2
Coagulation studies: PT/INR, aPTT, D-dimer for coagulopathy assessment 4
LDH: Typically elevated but non-specific 4
Lipid panel: Assess VLDL (elevated) and HDL (low) 4
Electrolytes: Check for hyponatremia and hypoproteinemia 4
Specialized Immunologic and Genetic Testing
Flow Cytometry
NK/T-cell degranulation assay (CD107a expression): Detects cytotoxic granule release defects in primary HLH 4
Perforin expression: Decreased or absent in familial HLH type 2 4
SAP protein expression: Decreased or absent in X-linked lymphoproliferative syndrome type 1 (XLP1) 4
Genetic Testing
- Next-generation sequencing panel: Test for mutations in PRF1, UNC13D, STX11, STXBP2, RAB27A, LYST, SH2D1A, BIRC4, and other HLH-associated genes 4, 5
CNS Evaluation (When Indicated)
- Cerebrospinal fluid analysis: Check for pleocytosis (mononuclear cells), elevated protein, and hemophagocytosis 4
- Strongly recommended in patients with neurological signs/symptoms or abnormal CSF parameters 4
Malignancy Screening
Peripheral blood smear and bone marrow: Screen for blasts and malignant cells 4
Imaging: Chest X-ray, abdominal ultrasound/CT, and lymph node imaging to exclude occult lymphoma 4
Tissue biopsy: Of suspicious lymph nodes or cutaneous lesions 4
Alternative Diagnostic Tool: HScore
For adults with suspected secondary HLH, the HScore provides a validated alternative scoring system that includes: known immunosuppression (0-18 points), temperature (0-49 points), organomegaly (0-38 points), number of cytopenias (0-34 points), ferritin level (0-50 points), triglyceride level (0-64 points), fibrinogen level (0-30 points), AST level (0-19 points), and hemophagocytosis on bone marrow (0-35 points) 4, 3
Critical Pitfalls to Avoid
Do not wait for all 5 criteria to be met before initiating treatment if clinical suspicion is high, as HLH is rapidly fatal without intervention 3
Hemophagocytosis may be absent initially and should not exclude the diagnosis; serial bone marrow examinations or tissue from other organs may be necessary 4, 3
Ferritin elevation must be distinguished from transfusion-related iron overload in malignancy contexts 4
The HLH-2004 criteria were developed for children and are not formally validated in adults, though they remain the standard diagnostic approach 4, 3
Many of these laboratory abnormalities can be caused by the underlying malignancy itself rather than HLH, requiring clinical judgment about whether the combination, extent, and progression are unusual and unexplained 4