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Test ID: PLINK Paroxysmal Nocturnal Hemoglobinuria, PI-Linked Antigen, Blood

Reporting Name

PNH, PI-Linked AG, B

Useful For

Screening for and confirming the diagnosis of paroxysmal nocturnal hemoglobinuria (PNH)


Monitoring patients with PNH

Specimen Type

Whole blood

Specimen Required

Specimen must arrive within 72 hours of collection.



Preferred: Yellow top (ACD)

Acceptable: Lavender top (EDTA)

Specimen Volume: 2.6 mL

Collection Instructions: Send whole blood specimen in original tube. Do not aliquot.

Specimen Minimum Volume

1 mL

Specimen Stability Information

Specimen Type Temperature Time Special Container
Whole blood Ambient (preferred) 72 hours
  Refrigerated  72 hours

Reference Values

An interpretive report will be provided. 



PNH RBC-Partial Antigen loss: 0.00-0.99%

PNH RBC-Complete Antigen loss: 0.00-0.01%

PNH Granulocytes: 0.00-0.01%

PNH Monocytes: 0.00-0.05%

Day(s) Performed

Monday through Friday

Test Classification

This test was developed using an analyte specific reagent. Its performance characteristics were determined by Mayo Clinic in a manner consistent with CLIA requirements. This test has not been cleared or approved by the US Food and Drug Administration.

CPT Code Information

88184-Flow cytometry, RBC x 1

88184-Flow cytometry, WBC x 1

88185-Flow cytometry, additional marker (each), RBC x 1

88185-Flow cytometry, additional marker (each), WBC x 6

88188-Flow Cytometry Interpretation, 9-15 Markers x 1

LOINC Code Information

Test ID Test Order Name Order LOINC Value
PLINK PNH, PI-Linked AG, B 90735-2


Result ID Test Result Name Result LOINC Value
CK079 Interpretation 90739-4
CK080 PNH RBC-Partial Ag Loss 33662-8
CK081 PNH RBC-Complete Ag Loss 90738-6
CK082 PNH Granulocytes 90737-8
CK083 PNH Monocytes 90736-0


Individuals with paroxysmal nocturnal hemoglobinuria (PNH) have absent or decreased expression of all the glycosylphosphatidylinositol (GPI)-linked antigens and fluorescent aerolysin (FLAER) on peripheral blood cells derived from the PNH clone.


Recent data showed that small PNH clones can be detected in a relatively high percentage of cases of aplastic anemia and myelodysplastic syndrome. While the significance of this finding is still uncertain, it appears that these patients may benefit from immunosuppressive therapy.


This test incorporates a sophisticated technique of separating different cell populations using gating on antigen-positive cells, as well as the sensitivity to enable detection of small PNH clones. In addition, this test detects a partial loss of CD59 on type II red blood cells (RBC). Patients with large proportion of type II RBC are unlikely to show high levels of hemolysis, unlike patients with complete loss of GPI-linked proteins (predominantly type III cells). While PNH is a disorder of hematopoietic stem cells and all lineages are affected, the percentage of affected cells can differ between lineages, most commonly due to hemolysis and/or transfusion.


Individuals without PNH have normal expression of FLAER (neutrophils and monocytes) and normal expression of all GPI-linked antigens-CD14 (monocytes), CD16 (neutrophils and NK cells), CD24 (neutrophils), and CD59 (RBC).

Clinical Reference

1. Richards SJ, Hill A, Hillman P: Recent advances in the diagnosis, monitoring and management of patients with paroxysmal nocturnal hemoglobinuria. Cytometry B Clin Cytom. 2007 Sep;72(5):291-298

2. Sutherland DR, Illingworth A, Marinov I, et al: ICCS/ESCCA consensus guidelines to detect GPI-deficient cells in paroxysmal nocturnal hemoglobinuria (PNH) and related disorders part 2 - reagent selection and assay optimization for high-sensitivity testing. Cytometry B Clin Cytom. 2018 Jan;94(1):23-48. doi: 10.1002/cyto.b.21610

3. Illingworth A, Marinov I, Sutherland DR, Wagner-Ballon O, DelVecchio L: ICCS/ESCCA consensus guidelines to detect GPI-deficient cells in paroxysmal nocturnal hemoglobinuria (PNH) and related disorders part 3 - data analysis, reporting and case studies. Cytometry B Clin Cytom. 2018 Jan;94(1):49-66. doi: 10.1002/cyto.b.21609

4. Oldaker T, Whitby L, Saber M, Holden J, Wallace PK, Litwin V: ICCS/ESCCA consensus guidelines to detect GPI-deficient cells in paroxysmal nocturnal hemoglobinuria (PNH) and related disorders part 4 - assay validation and quality assurance. Cytometry B Clin Cytom. 2018 Jan;94(1):67-81. doi: 10.1002/cyto.b.21615

5. Dezern AE, Borowitz MJ: ICCS/ESCCA consensus guidelines to detect GPI-deficient cells in paroxysmal nocturnal hemoglobinuria (PNH) and related disorders part 1 - clinical utility. Cytometry B Clin Cytom. 2018 Jan;94(1):16-22. doi: 10.1002/cyto.b.21608

6. Illingworth AJ, Marinov I, Sutherland DR: Sensitive and accurate identification of PNH clones based on ICCS/ESCCA PNH Consensus Guidelines-A summary. Int J Lab Hematol. 2019 May;41 Suppl 1:73-81. doi: 10.1111/ijlh.13011

7. Seth N, Mahajan V, Kedia S, Sutar A, Sehgal K. Utility of FLAER and CD157 in a five-color single-tube high sensitivity assay, for diagnosis of Paroxysmal Nocturnal Hemoglobinuria (PNH)-A standalone flow cytometry laboratory experience. Int J Lab Hematol. 2021 Apr;43(2):259-265. doi: 10.1111/ijlh.13366

8. Payne D, Johansson U, Bloxham D, et al: Inter-laboratory validation of a harmonized PNH flow cytometry assay. Cytometry B Clin Cytom. 2018 Sep;94(5):580-587. doi: 10.1002/cyto.b.21726

9. Sutherland DR, Ortiz F, Quest G, et al: High-sensitivity 5-, 6-, and 7-color PNH WBC assays for both Canto II and Navios platforms. Cytometry B Clin Cytom. 2018 Jul;94(4):637-651. doi: 10.1002/cyto.b.21626

Report Available

1 to 2 days

Clinical Information

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired hematologic disorder characterized by nocturnal hemoglobinuria, chronic hemolytic anemia, thrombosis, pancytopenia, and, in some patients, acute or chronic myeloid malignancies.


PNH appears to be a hematopoietic stem cell disorder that affects erythroid, granulocytic, and megakaryocytic cell lines. The abnormal cells in PNH have been shown to lack glycosylphosphatidylinositol (GPI)-linked proteins in erythroid, granulocytic, megakaryocytic, and, in some instances, lymphoid cells. Variants in the phosphatidylinositol glycan A gene, PIGA, have been identified consistently in patients with PNH, thus confirming the biological defect in this disorder.


A flow cytometric-based assay can detect the presence or absence of these GPI-linked proteins in granulocytes, monocytes, erythrocytes, and lymphocytes, thus avoiding the problems associated with red blood cell (RBC)-based diagnostic methods (Ham test) in which recent hemolytic episodes or recent transfusions can give false-negative results. A partial list of known GPI-linked proteins includes CD14, CD16, CD24, CD55, CD56, CD58, CD59, C8-binding protein, alkaline phosphatase, acetylcholine esterase, and a variety of high frequency human blood antigens. In addition, fluorescent aerolysin binds directly to the GPI anchor and can be used to evaluate the expression of the GPI linkage.


In-house studies, as well as others in the literature, have shown that flow cytometry-based assays will detect all Ham-positive PNH cases, as well as some Ham-negative PNH cases. This assay replaces the sugar water test and the Ham test for the evaluation of patients with possible PNH.


Patients with PNH should be transfused with ABO-specific RBCs, which do not need to be washed. If, for some reason, they need to receive non-ABO type-specific (type O) cells, these RBC units should be washed. Since recipient antibodies to granulocyte antigens can trigger hemolytic episodes in PNH, if they have such antibodies these patients should receive leukoreduced RBCs and platelets.

Method Name


Additional Tests

Test ID Reporting Name Available Separately Always Performed
FCIMS Flow Cytometry Interp, 9-15 Markers No, (Bill Only) Yes
Mayo Clinic Laboratories | Hematology Catalog Additional Information:

mml-benign-hematology-disorders, mml-acute-leukemia-myelodysplastic-syndromes