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HelpTEST ID PLINK Paroxysmal Nocturnal Hemoglobinuria, PI-Linked Antigen, Blood
Reporting Name
PNH, PI-Linked AG, BSpecimen Type
Whole bloodShipping Instructions
Specimen must arrive within 72 hours of collection.
Specimen Required
Container/Tube:
Preferred: Yellow top (ACD solution A or B)
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 |
|---|---|---|
| Whole blood | Ambient (preferred) | 72 hours |
| Refrigerated | 72 hours |
Method Name
Immunophenotyping
Reject Due To
| Gross hemolysis | Reject |
| Fully clotted | Reject |
Reference Values
An interpretive report will be provided.
RED BLOOD CELLS:
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 Saturday
Report Available
1 to 3 daysSpecimen Retention Time
14 daysPerforming Laboratory
Mayo Clinic Laboratories in Rochester
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
Forms
If not ordering electronically, complete, print, and send 1 of the following forms with the specimen:
Useful For
Screening for and confirming the diagnosis of paroxysmal nocturnal hemoglobinuria (PNH)
Monitoring patients with PNH
Cautions
The sugar water test and the Ham test are no longer recommended for the evaluation of patients with possible paroxysmal nocturnal hemoglobinuria.
Recent transfusion can decrease the sensitivity of this test and interfere with accuracy.
Method Description
Flow cytometric immunophenotyping of peripheral blood (white blood cells [WBCs] and red blood cells [RBCs]) is performed using the following antibodies:
RBC: CD235a, CD59
WBC: CD14, CD15, CD16, CD24, CD33, CD45, and FLAER
This assay evaluates the presence or absence of glycosylphosphatidylinositol (GPI)-linked proteins using monoclonal antibodies directed against CD235, CD33, and CD15 to isolate different cell lineages. GPI-linked proteins that are checked within different lineages include CD14 for monocytes, CDs 16 and 24 for granulocytes, and CD59 for RBC. Fluorescent aerolysin, a fluorescently labeled inactive variant of the protein aerolysin, binds selectively to GPI anchors and is evaluated for presence or absence of expression on WBCs. In addition, this test will detect a partial loss of CD59 on RBCs (type II RBC).
Individuals without paroxysmal nocturnal hemoglobinuria have normal expression of all GPI-linked antigens on peripheral blood and leukocytes and erythrocytes.(Devalet B, Mullier F, Chatelain B, Dogne JM, Chatelain C. Pathophysiology, diagnosis, and treatment of paroxysmal nocturnal hemoglobinuria: a review. Eur J Haematol. 2015;95(3):190-198. doi:10.1111/ejh.12543)
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.
Paroxysmal nocturnal hemoglobinuria 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.
Additional Tests
| Test ID | Reporting Name | Available Separately | Always Performed |
|---|---|---|---|
| FCIMS | Flow Cytometry Interp, 9-15 Markers | No, (Bill Only) | Yes |