This study investigates PCR analysis of immunoglobulin heavy chain (IgH) and T cell receptor (TCR) gene rearrangements on paraffin-embedded tissue sections and bone marrow aspirates of patients suspected to have lymphoproliferative disorders but with inconclusive diagnosis in histopathological examination. 130 samples of patients with inconclusive immunohistochemistry results were evaluated for clonal rearrangement of IgH and TCR genes. Based on histopathology examination, the patients were divided into three groups: the first group without any definite diagnosis of lymphoproliferative disorders (60 cases, 46.2 %), the second group suspected to have a lymphoproliferative disorder but in favor of benign disorders (19 cases, 14.6 %) and the third group suspect to lymphoproliferative disorders but relatively in favor of malignant disorders (51 cases, 39.2 %). After DNA extraction and quality control, semi-nested PCR was performed using consensus primers for amplification of TCR-c and CDR-3 regions of IgH genes. PCR products were analyzed after heteroduplex analysis using polyacrylamide gel electrophoresis, and were subject to silver staining. Totally, in over half of the cases (55.4 %), a monoclonal pattern was found in IgH or TCR-c genes rearrangements. Monoclonal IgH gene rearrangement was detected in 48.1 % of patients, whereas monoclonal TCR-c gene rearrangement was found in 33.6 % of them, which was not statistically significant (P = 0.008). Only in 32 patients (24.6 %) were the results of TCR-c and IgH gene rearrangements consistent with respect to the presence (2.3 %) or absence (22.3 %) of monoclonality. Finally, PCR analysis of TCR-c and IgH gene rearrangements led to definite diagnosis in 105 patients (80.8 %), and only 25 cases (19.2 %) remained inconclusive. Our results emphasize the usefulness of gene rearrangement study in cases without a definite diagnosis in immunohistochemistry studies. Multiple PCR analysis results when combined with patient's clinical course and immunohistochemistry can lead to early diagnosis and subsequent therapy.

Since paroxysmal nocturnal haemoglobinuria (PNH) was first described in 1881, the diagnosis and follow-up patients diagnosed with the illness has remained an area of concern, with several different techniques of varying sensitivity having been described in the literature for both the diagnosis and monitoring treatment of the disease. PNH is a rare and life-threatening disease that manifests symptoms of haemolytic anaemia.Hence, a quick and reliable technique for precise diagnosis would be crucial. PNH patients who have previously been diagnosed with aplastic anaemia or myelodysplastic syndrome carry small PNH clones and for more than a century traditional method with low sensitivity was used for such patients. In 2010, the International Clinical Cytometry Society described a highly sensitive method for detection and quantification of different types of PNH clones using multi-colour flow cytometry. In this method, a threecolour flow cytometer is essential to detect PNH affected cells amongst monocytes and granulocytes. This started a new era in the diagnosis of patients who carry small clones of PNH cells. Before this, flow cytometric analysis was used only for detection of PNH cells amongst erythrocytes. By using flow cytometry instruments with more light sources, the sensitivity of detection and quantification of PNH clones would be augmented. However, standardisation and crosstalk compensation would be the most concerning issue. For the first time in Iran, we set up and standardised multi-colour flow cytometry technique to detect PNH cells in erythrocytes and leukocytes at Payvand medical laboratory.