Individuals with paroxysmal nocturnal hemoglobinuria (PNH) often present with nonspecific symptoms that may overlap with other hematologic and renal conditions, which can result in a delayed diagnosis. The timely and accurate diagnosis of PNH can lead to improved treatment and patient outcomes.

As far as formulating a differential diagnosis of PNH, the most common initial step is going to be performing a workup for anemia. When a patient has anemia, it typically falls into 3 “buckets”: blood loss, a production problem, or hemolysis. Hemolysis is actually the least common of the 3 buckets, so it is not something that I routinely look for in every patient with anemia, unless there are risk factors that you can identify from the initial patient history. Other signs or symptoms that should make you think about PNH up front would include the presence of hemolysis and thrombosis at unusual sites, such as the cerebral, hepatic, portal, and splenic veins.

A patient with classic PNH generally has a hemoglobin level of under 10 g/dL, fatigue, shortness of breath, a history of unprovoked blood clots, and smooth muscle dysfunction. The reticulocyte is my best indicator to distinguish issues with red blood cell destruction from production. PNH is associated with increased red blood cell destruction, not a production problem. If the patient’s reticulocyte count is below a normal level, then there is something preventing production, such as aplastic anemia, myelodysplastic syndrome, or a nutritional deficiency, and it may be necessary to check the bone marrow for a possible secondary pathology. Reticulocytosis in a patient’s laboratory results would suggest a hemolytic anemia. This tells me that they are anemic but are producing red blood cells adequately, and if there is also no evidence of bleeding, then it is most likely due to a destructive process or hemolysis.

Once you go down that path, you then perform additional tests to confirm the cause of hemolysis, including elevated serum lactate dehydrogenase, elevated bilirubin, decreased haptoglobin, and peripheral blood smear, which can help narrow down the reasons for hemolysis. The hemoglobin A_1C test can also be a marker for hemolysis. If a patient’s hemoglobin A_1C level is below normal and there is no evidence of long-term hypoglycemia, that is suspicious for hemolysis. Once you confirm hemolysis, we use the direct antiglobulin (Coombs) test to narrow it down further. If the patient has a positive Coombs test result, that is considered antibody-mediated hemolysis. If they have a negative Coombs test result, then, in addition to PNH, it could be many other nonantibody-mediated things, such as a mechanical heart valve, medications, an infection, or a thrombotic microangiopathy.

Once the common causes of hemolysis are ruled out and the patient’s Coombs test is negative, then the classic test to confirm PNH is flow cytometry. We look for the GPI-anchored proteins CD55 and CD59. We also use fluorescently labeled aerolysin, which more broadly detects GPI itself. If those proteins are not detected, that is confirmation that the patient has PNH.