Flow cytometry is the most sensitive and specific diagnostic modality for paroxysmal nocturnal hemoglobinuria (PNH) clone assessment in PNH and other bone marrow failure states. Hemolysis 1. REQUEST FLOW CYTOMETRY ON TISSUE FOR: Suspected lymphoma (past history of lymphoma, lymphadenopathy, radiology/imaging suggests lymphoma) Flow is of no value in Hodgkin lymphomas TISSUE SAMPLE REQUIREMENTS Separate fresh tissue specimen in sterile pot or in tissue preservative (RPMI, pink liquid - preferred if sending after hours) Borowitz MJ, Craig FE, Digiuseppe JA, et al. Symptoms include dysphagia, lethargy, renal failure, anemia, hemoglobinuria, male impotence, pulmonary . It has not been cleared or approved by the U.S. Food and Drug Administration. Board certified hematopathologists trained on each available platform review and interpret flow cytometry results, including assessments of minimal residual disease by flow cytometry for leukemias, lymphomas and plasma cell neoplasms. . In clinical flow cytometry, monoclonal antibodies should be validated in the context of the assay as part of the assay validation procedure. 2018 Sep;94(5):580-587. doi: 10. . The extent of the defect is more related to haemolysis than the haematopoietic deficiency. Paroxysmal nocturnal hemoglobinuria (PNH) affects your bone marrow, where your body makes new blood cells. PNH test by Flow Cytometry, Blood - Canterbury Health Laboratories. Flow cytometric immunophenotyping is performed to detect the presence or absence of GPI-linked proteins on granulocytes, monocytes, and erythrocytes, using fluorescently labeled monoclonal antibodies or FLAER. Cytometry B Clin Cytom. Advanced 10-color flow cytometry and rapid turnaround times. The flow cytometric assay evaluates for a loss of expression of the following GPI-linked antigens: CD59 on red cells, CD14 and FLAER on monocytes, and CD24 and FLAER on granulocytes. 4-6 single somatic mutation of the piga gene in Flow cytometry has been used in diagnosing other disorders such as paroxysmal nocturnal hemoglobinuria (PNH, a stem cell disorder), 15 feto-maternal hemorrhaging (seen in hemolytic disease of the newborn), 16 various newborn immunodefiencies, 2,8 abnormal granulocyte function activities (patients with chronic granulomatous disease 17), and . Sugar Water PNH Screen. . 2. Specimen Preparation Transport 4 mL whole blood. This is generally done on a bone marrow sample. Refer to ACS 'Guidelines for Clinical Flow Cytometry Laboratory Practice' for additional A total of 101 samples included 23 PNH, 46 aplastic anemia (AA), seven myelodysplastic syndrome (MDS) cases, and 25 normal controls. Specific groupings of these antigens are normally present on or within WBCs and are unique to specific cell types and stages of cell . Regulatory Links. 9. Flow cytometric GPI-AP analysis was performed in 803 patients suffering from PNH or suspected GPI-deficient populations. The results of PNH testing by flow cytometry are usually reported as percentage of type II and III PNH cells from the total gated neutrophils, monocytes and red blood cells. Approach Considerations In addition to a complete blood cell count, the principal studies used to establish the diagnosis of paroxysmal nocturnal hemoglobinuria (PNH) are flow cytometry of. Granulocytes (red) in peripheral blood derived from PNH clone are . Last-updated: 29/12/21. Epic Code LAB2111417 Paroxysmal Nocturnal Hemoglobinuria (PNH) by Flow Cytometry Test Name Alias. This assay is a rapid flow cytometry screening test for the presence of the SAP protein . The reason for flow cytometric analysis as reported by the treating physician were the following diagnoses or suspected diagnoses: PNH in 67 patients, hemolysis in 77 patients, aplastic anaemia or trilineage cytopenia in 291 patients, MDS in 44 patients, single . In the last issue of the eNewsletter Marc Golightly described the DHR test for chronic granulomatous disease and mentioned how this flow cytometric test has replaced the Nitroblue tetrazolium dye reduction test (NBT). 1 2 3 The variant leads to a lack of glycosylphosphatidylinositol (GPI)-anchored proteins on the surface of blood cells, which in turn leads to an inappropriate immune response to, and hemolysis of . Search. Bloxham D, et al: Inter-laboratory validation of a harmonized PNH flow cytometry assay. During the process, a sample of cells or particles is suspended in fluid and injected into a flow cytometer machine. paroxysmal nocturnal hemoglobinuria (PNH) clones by flow cytometry. The flow cytometry test measures how many of your blood cells are missing a protein . PNH PAN Methodology Quantitative Flow Cytometry Performed Sun-Sat Reported 1-3 days New York DOH Approval Status This test is New York DOH approved. A Standard is the minimum requirement for a procedure, method, staffing resource or . These antigens are protein structures found on or within WBCs. This test tells your doctor if any proteins are missing from the surface of blood cells. Hillmen P, Young NS, Schubert . Patients. For analysis, RBCs were identified by light-scatter properties. This issue of the eNewsletter provides a few other examples. Bloxham D, et al: Inter-laboratory validation of a harmonized PNH flow cytometry assay. Cytometry B Clin Cytom. It is highly recommended to use well described monoclonal antibodies derived from clones described by the Human Leukocyte Differentiation Antigen (HLDA) Workshops. The gold standard for confirming the presence of PNH is a flow cytometry test. Send whole blood specimen at ambient temperature. This test code is for non-New York patient testing. ( 7) Flow cytometry (FCM) is the gold standard for PNH diagnosis and monitoring, even in patients with small PNH clones. Today, it is fair to say that outside of research settings the value of PNH testing by flow cytometry is largely limited to diagnosis, and that a small panel of markersincluding some directed against GPI-linked granulocyte antigens is best used to make sure that all patients with the disease are identified. Paroxysmal nocturnal hemoglobinuria (PNH). This blood test can separate and analyze various components of blood. Specimen must be received within 24 hours of collection. Along with high sensitivity and specificity, the technique is quick to perform and provides qualitative and quantitative analysis of GPI-anchored proteins. Flow cytometry was performed using CD55, CD59, and stain-lyse-wash method, and . A flow cytometry test is considered the gold standard for diagnosing PNH. This tool counts and examines them. Flow cytometry is a technology that provides rapid measurements of multiple characteristics of cells suspended in a moving fluid stream as it passes through multiple laser beams. Sutherland DR, Ortiz F, Quest G, et al: High . Listen to staff scientists as they share clinical flow cytometry testing capabilities in these short videos. The flow cytometric assay evaluates for a loss of expression of the following GPI-linked antigens: CD59 on red cells, CD14 and FLAER on monocytes, and CD24 and FLAER on granulocytes. feto-maternal hemorrhage. Processing. The Dahl-Chase Flow Cytometry division is a client-oriented, cutting-edge, full-service flow cytometry laboratory with extensive experience in PNH (paroxysmal nocturnal hemoglobinuria) testing. . A flow cytometer evaluates the physical and/or chemical characteristics of single cells as the cells pass . Flow cytometry is the gold standard in diagnosis of paroxysmal nocturnal hemoglobinuria (PNH) by detecting the absence of glycol-phosphatidyl inositol (GPI)linked protein expression on granulocyte and monocyte surfaces. Minor population of CD55-CD59- blood cells predicts response to immunosuppressive therapy and prognosis in patients with aplastic anemia. Paroxysmal nocturnal hemoglobinuria (PNH) is a rare, potentially life-threatening acquired stem cell disorder caused by a variant in the PIGA gene. FLAER is a new type of flow cytometry test that is also used. It reveals whether two key proteins, CD55 and CD59, are missing from the surface of blood cells. Licenses & Accreditation Specialty Testing . Laboratory Test Guide. Sample type guide. Identification of large PNH clone by flow cytometry. The assay can detect as little as 0.01% GPI-deficient cells in each cell lineage. EDTA-anticoagulated whole blood was stained with anti-CD 55-Phycoerythrin (PE) and anti-CD 59-PE.A minimum of 10,000 events were acquired. Flow Cytometry for Hematologic Malignancies Diagnosis Comprehensive panels are useful for the diagnosis of leukemia, lymphoma, and plasma cell neoplasms. Test Description FLAER(A488), CD24, CD14, CD33, CD15, Near-IR, CD45 . Specimen Type. Diagnosis of PNH Conventional laboratory tests for the diagnosis of PNH include the sugar water test and the Ham's acid hemolysis test . Message from the Editor: Flow cytometric procedures have replaced several well established clinical laboratory tests. 88184 First marker flow cytometry, technical; 88185 Each additional marker, flow cytometry technical; 88187 Flow cytometry interpretation, 2 - 8 markers; 88188 Flow cytometry interpretation, 9 - 15 markers; 88189 Flow cytometry interpretation, 16 or more markers; 88319 Myeloperoxidase or non-specific esterase stain (for acute leukemia) Flow cytometry (FCM) is a complex process to examine blood, body fluids, CSF, bone marrow, lymph node, tonsil, spleen and . PNH Treatment Antibodies to CD55 and CD59 are specific for decay-accelerating factor and membrane-inhibitor of reactive lysis, respectively, and can be analyzed by flow cytometry to make a definitive diagnosis of PNH. www.cap.org . References Reference Range. PNH Panel CD14, CD15, CD24, CD45, CD59, CD64, CD235a, FLAER . Blood 2006; 107: 1308-1314. The first test is comprised of a flow cytometry analysis, which helps to determine if any blood cell lacks the GPI-anchored proteins. Back to test list. . Specimen Required Patient Preparation Collect Lavender (EDTA), pink (K 2 EDTA), or green (sodium or lithium heparin). . It can measure certain proteins associated with PNH to diagnose the disease. C1.2(i) The test should be performed within 48 hours of collection (3) for white blood cells (WBC) but up to 72 hours (EDTA) may be . The PNH blood test will confirm if the disease is present. What is does is measure the percentage of blood cells in the body that are affected by the disease. CLIA Certification (2021 to 2023) CAP Certification Based on our review of the literature and our clinical experience, our current recommendations for diagnosis of PNH via flow cytometry are as follows: The specimen should consist of 5 mL of EDTA-anticoagulated whole blood. The Laboratory Accreditation Program inspects a variety of laboratory settings from complex university medical centers to physician office laboratories, and covers a complete array of disciplines and testing procedures. Specimen Collection Handling Procedure; Common ICD-10 Codes for Flow Cytometry; Services; PNH with FLAER (High Sensitivity) Test Details Methodology Flow Cytometry (FC) Assay Category This test was developed and its analytical performance characteristics have been determined by Quest Diagnostics. Flow cytometry is used for immunophenotyping of a variety of specimens, including whole blood, bone marrow, serous cavity fluids, cerebrospinal fluid, urine, and solid tissues. See the Special Instructions Legend page for a full list of special instructions. Absence or reduced expression. J. The flow cytometric (FCM) demonstration of the absence of GPI- linked molecules in a sizable fraction of periph- eral blood red cells, neutrophils and monocytes is the cornerstone of the diagnostic process of PNH (15-17). Reference Range(s) See Laboratory Report. Sugar Water Test, Ham-Crosby, Blood. Performance. 2018 Sep;94(5):580-587. doi: 10. . FLO.23706 Record Retention - Flow Cytometry Phase II Flow cytometry data for evaluation of hematolymphoid neoplasias, PNH, and congenital immunodeficiency evaluations are retained for at least 10 years. Test Method Flow Cytometry Disease / Condition Paroxysmal Nocturnal Hemoglobinuria Disease Abbreviation PNH CPT Codes 88184x1 88185x6 88187. Symptoms. For New York patient testing, use test code 37340. Approximately 10,000 cells can be analyzed and processed by a computer in less than one minute. Flow cytometry antibodies, assays and reagents This procedure is a flow cytometric assay that uses whole blood to screen for abnormalities in the expression of CD40-Ligand, CD40-Ig FP (Fusion Protein) and ICOS (inducible co-stimulator) on the surface of in vitro activated CD4+ T cells. It can also be used to check the health of the bone marrow after a transplant or after chemotherapy. If there is going to be a delay of more than 6 hours, the specimen should be stored at 4C. Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired clonal stem cell disorder that leads to intravas- Paroxysmal Nocturnal Hemoglobinuria by Flow Cytometry Toggle Page Navigation View pages within this section . Characteristics that can be measured include cell size, cytoplasmic complexity, DNA or RNA content, and a wide range of membrane-bound and intracellular proteins. PNH cells are missing some or all of two proteins on their surface. Flow Cytometry is a highly complex process by which blood, body fluids, bone marrow and tissue can be examined. Bone Marrow Test Contact laboratory for advice on sample taking. See the Sample Requirements page for an explanation of all the sample requirements. (Min: 4 mL) The use of peripheral blood and fine needle aspirate material avoids more invasive procedures for diagnosis. Bloxham D, et al: Inter-laboratory validation of a harmonized PNH flow cytometry assay. What Is the PNH Blood Test? This innovative laboratory was pivotal in the development of a very sensitive method for the detection of minor PNH clones that is the national standard . This can help to determine the cause of anemia. PNH by Flow Cytometry Method. The second test is carried out to analyze markers in blood for establishing the presence of hemolysis. The instrument is now used not only for research but also for routine clinical activities [ 1, 2, 3 ]. so a doctor will recommend this procedure only if you have severe PNH that hasn't responded well to other treatments. Specimen Collection and Handling. Three types of clinical presenta- tion of PNH are schematically considered: the classical form, associated to overt hemolysis Ordering & Collection. PNH |CD59 | P1 linked antigen | P1 Anchor | GPI linked antigen | Flaer | High Sensitivity PNH | 915. . This test is good for early detection of PNH clones in AA patients at initial diagnosis and in long term survivors. The flow cytometry test is used to count the number of reticulocytes (immature red blood cells) in the bone marrow. It is the method of choice for the detection of very small PNH clones in subclinical PNH that often accompanies Routine lymphocyte subset and CD34+ enumeration data are retained for at least two years. Date of collection and clinical information (to include diagnosis/suspected diagnosis) are required on request form for processing (specify PNH testing). It provides important immunophenotypic and DNA cycle information, of both diagnostic and prognostic interest in hemopathology, cytopathology and general surgical pathology. Cytometry B Clin Cytom. Methods: FCM provides us very quick assessment of cell surface antigens, DNA content and intracellular proteins. Sugimori C, Chuhjo, Feng X, et al. While flow cytometry is the method of choice for identifying cells deficient in GPI-linked proteins and is, therefore, necessary for the diagnosis of PNH, to date there has not been an attempt to standardize the methodology used to identify these cells. The particles are then passed by an electronic detection apparatus. Although the flow cytometry method is technically superior to the Kleihauer-Betke test, cost, instrument availability, and stat access may limit its practical utility. Cytometry Part B 2012; 82B: 195-208. Flow cytometry is a lab test used to analyze characteristics of cells or particles. The results will either be positive or negative. PNH patients have decreased expression of a protein called glycophosphatidylinositol (GPI) anchor proteins on red blood cells.. 2010;78 (4):211-230. The state-of-the-art laboratory test is flow cytometry of the patient's blood to detect CD59 (MIRL), a glycoprotein, and CD55 (DAF) in regulation of complement action. Describes the specimen type validated for testing . flow cytometry has become the most sensitive diagnostic tool for paroxysmal nocturnal hemoglobinuria (pnh) 1-3 in use since this rare hematopoietic stem cell disorder is characterized by the absence and/or reduced expression of glycophosphatidylinositol (gpi)-anchored proteins in both wbcs and rbcs. References They can also be used for post-treatment follow-up or evaluation of myeloid maturation. . Flow cytometry suspends microscopic particles in a stream of fluid. 1. Flow cytometry has become the gold standard method for PNH clone detection due to its high sensitivity and specificity and due to the ability to examine multiple GPI- linked proteins on red and white blood cell surface. 3. Flow Cytometry We are committed to accelerating your science by providing a comprehensive suite of solutions for the analysis of cells and their function by delivering you our flagship flow cytometry products designed to deliver high-performance results and save you time. Interpretation. Flow cytometry (FCM) is the method by which the various characteristics of individual particle or cells are studied. Fabric face coverings are no longer acceptable. 2018 Sep;94(5):580-587. doi: 10.1002/cyto.b.21726. The assay can detect as little as 0.01% GPI-deficient cells in each cell lineage. See our COVID-19 pages for detailed information about hospital visiting guidelines , COVID-19 tests and care in the . Cytometry B Clin Cytom. LIS Dept Code. The diagnosis is based on the Flow Cytometric (FCM) detection of peripheral blood cell clones lacking the su Lab Code. Flow cytometry is the most sensitive and specific diagnostic modality for paroxysmal nocturnal hemoglobinuria (PNH) clone assessment in PNH and other bone marrow failure states. Assays reporting numeric data are considered as semi-quantitative, therefore the post-analytical validation process should comprise confirmation of accuracy, specificity . Flow cytometry is a blood test used to diagnose PNH. negative for GPI-linked proteins and FLAER. The official name of this test is called the "high-sensitivity flow cytometry" test. Standard Leukemia/Lymphoma Panel - 24 markers Extended Leukemia/Lymphoma Panel - 31 markers Guidelines for the diagnosis and monitoring of paroxysmal nocturnal hemoglobinuria and related disorders by flow cytometry. Four color flowcytometry immunophenotypic analysis with CD55 and CD59 was used to detect PNH clones in RBCs. The advantage of flow cytometry is, that even in nonhemolysing patients, small PNH clones can be detected and the proportion of type III (complete deficiency), type II (partial deficiency) and type. LOINC Codes, Performing Laboratory PNH A more sensitive assay (0.004%) using a bacterial toxin-aerolysin which spares PNH cells Red cells show clearest delineation between phenotypes Assay both erythrocytes and granulocytes Small proportion (esp PNH/AA) may only be detected on granulocyte clones PNH Summary: Analysis of GPI-AP - highly specific test for PNH. However, the current assays are not optimized and require improvement, particularly in reducing background fluorescence and optimizing sensitivity and specificity. In both diseases it permits measuring the extent of the abnormal clone and its follow up. Immunophenotyping by flow cytometry is a laboratory method that detects the presence or absence of white blood cell (WBC) markers called antigens. Paroxysmal Nocturnal Hemoglobinuria (PNH) is a rare blood disorder characterized by chronic intravascular hemolysis, thromboses in unusual sites and cytopenias related to bone marrow failure. Hospital visiting guidelines updated 20 July 2022: Hospital visitors must wear a surgical/medical paper mask. A definitive diagnosis of PNH is available through flow cytometry. Flow cytometric immunophenotypic analysis is the method of choice to detect populations of GPI anchor-deficient cells used in the diagnosis of PNH and to monitor patients with an established diagnosis.1PNH erythrocyte clones may be divided into those with partial loss of CD59 (PNH Type II Cells) or complete loss of CD59 (PNH Type III Cells). PNHMET. at a minimum, an evaluation of a pnh result should include (1) pnh flow cytometry, which should be performed on peripheral blood with quantification of pnh neutrophils and pnh rbcs; (2) a bone marrow aspirate and biopsy, including cytogenetics (testing for somatic mutations associated with hematologic malignancies can also be helpful for These proteins are called CD55 and CD59. Alternative Name(s) Flow,Leukemia Panel,Flow Cytometry Leukemia/Lymphoma,Lymphoma Panel,Leukemia/Lymphoma Phenotyping Panel,Immunophenotyping. 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. 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. With . Organ Transplants . Specimen Type Collection. Flow cytometry analysis of peripheral blood is the current gold standard for laboratory detection of PNH.