Sehgal index: A new index and its comparison with other complete blood count-based indices for screening of beta thalassemia trait in a tertiary care hospital.

Introduction: Beta thalassemia trait (BTT) must be differentiated from iron defi ciency anemia to avoid unnecessary iron therapy and for the prevention of thalassemia major by genetic counseling. In a tertiary care hospital, it is vital that the screening tool is not only sensitive but also specifi c so as to be cost effective and save time. Aim: The aim of this study was to evaluate the new Sehgal index and compare it to existing complete blood count-based indices for the best combination of sensitivity and specifi city to predict BTT. Materials and Methods: Study was done in 2 phases - Phase 1: A retrospective analysis of 1022 consecutive high-performance liquid chromatography (HPLC) cases from July 2008 to June 2011. Phase 2: A prospective analysis of 973 consecutive HPLC cases from July 1, 2011 to June 10, 2013 was done to confi rm the results of Phase 1 and the applicability of the new Sehgal index. Results: Prevalence of BTT was 28.8% (294/1022) and 25.39% (247/973) in Phase 1 and Phase 2, respectively. Receiver operating characteristic-area under the curve and Youden index was highest for new Sehgal index, followed by Mentzers index <14. The prospective study shows results similar to those in Phase 1 confi rming the superiority of the above two indices. Conclusion: Sehgal index and Mentzers index <14 showed the best combination of sensitivity and specifi city in predicting BTT. The best indices or combination can be used as a “validated fl ag rule” in the analyzer middleware program in a hospital for identifying suspected cases of BTT.

Unusual immunophenotype of T-cell large granular lymphocytic leukemia: Report of two cases.

Large granular lymphocytes (LGL) leukemias are commonly of the T-cell or NKcell type. T-cell LGL leukemia is typically a disorder of mature CD3, CD8 and T-cell receptor TCR (TCR — T cell receptor)-αβ positive cytotoxic T-cells. Rare variants include TCRγδ+ variants and CD4+ TCRαβ+ cases. We report a case of each of these rare variants. An 83-year-old female presented with anemia and lymphocytosis with LGLs on peripheral smear. Six-color multiparametric fl owcytometric analysis showed expression of CD3, heterogeneous CD7, dim CD2 and TCRγδ and lacked expression of CD5, TCRαβ, CD56, CD4 and CD8. A fi nal diagnosis of TCRγδ+ T-cell LGL leukemia was made. Differentiation between TCRγδ+ T-cell LGL leukemia and other γδ+ T-cell malignancies is of utmost importance due to the indolent nature of the former as compared to the highly aggressive behavior of the latter. An 85-year-old male diagnosed with liposarcoma was identifi ed to have lymphocytosis during preoperative evaluation. Peripheral smear showed presence of LGLs. Flowcytometric immunophenotyping showed expression of TCRαβ, CD3, CD2, CD5, CD4, dim CD8, CD56 with aberrant loss of CD7 expression. Vβ repertoire analysis by fl owcytometry showed 97% cells with Vβ14 clonality. A fi nal diagnosis of TCRαβ+ CD4+ T-cell LGL leukemia was made. CD4+ T-cell large granular lymphocytic leukemias have an indolent, less aggressive course when compared to their CD8+ counterparts and are not necessarily associated with cytopenias. However, their association with secondary neoplasia (29% of the cases) warrants a high degree of suspicion in the diagnosis as also noted in the index case. Use of a wide panel of antibodies and newer modalities such as Vβ repertoire analysis helps in accurate subtyping of LGL leukemia

Reference range evaluation of complete blood count parameters with emphasis on newer research parameters on the complete blood count analyzer Sysmex XE-2100.

Since the advent of automation in the fi eld of hematological cell counters there has been a constant refi nement of the technology and increase in the number of newer parameters available on CBC analysers. Many novel parameters are being put into routine clinical use and both clinical evaluation and monitoring critically depend on knowledge of laboratory reference ranges. Here, we present reference interval for the Sysmex XE-2100, with emphasis on the novel or newer research parameters. Blood samples from a total of 122 clinically asymptomatic and apparently healthy subjects were evaluated and a fi nal of 100 subjects (54-M, 46-F) were included in the study. A broad spectrum of parameters available with the analyser was assessed and reference ranges for the same evaluated.

Comparison of platelet counts by sysmex XE 2100 and LH-750 with the international flow reference method in thrombocytopenic patients.

Background: There are several methods for counting platelets, of which the international fl ow reference method (IRM) is considered to be the gold standard. We compared the platelet count given by this method to the count given by automated analyzers using other methods, such as optical fl uorescence and impedance. Aims: The aim of this study is to compare the platelet counts obtained by Sysmex XE 2100 by Impedance (Sysmex-I), optical fl orescence (Sysmex-O) and reported (Sysmex-R) based on the switching algorithm and LH-750 by Impedance (LH-750) with the IRM in thrombocytopenic blood samples. To calculate the sensitivity, specifi city, positive predictive value (PPV) and negative predictive value (NPV) of various technologies at the clinically relevant transfusion thresholds of 10 × 109/l and 20 × 109/l. Materials and Methods: A total of 118 blood samples with platelet count of <50 × 109/l were selected for the study. Platelet counts of all samples were analyzed by all methods using the Sysmex analyzer, LH-750 and IRM in parallel within 6 h of collection. Statistical Analysis Used: Pearson correlation, bland Altman analysis, sensitivity and specifi city, PPV and NPV. Results and Conclusions: Sysmex-R had the least Bias and 95% limits of agreement (95%LA) range and thus correlated best with IRM values. LH-750 had a higher Bias compared to Sysmex-O and Sysmex-R, but a strikingly similar 95% LA ensures similar results in all three methods. In fact, in the oncology subset, it had the narrowest 95% LA, which made it the best performer in this subgroup. Of the three Sysmex results, Sysmex-I had the highest bias, widest 95% LA and highest potential risk of over transfusion. Hence, Sysmex-R and LH-750 were found to be reliable tools for estimation of platelet count in thrombocytopenic patients.

Leukemic phase of anaplastic lymphoma kinase positive, anaplastic large cell lymphoma.

Anaplastic large cell lymphoma (ALCL) is a distinct type of CD30+ T/null-cell non-Hodgkin's lymphoma that frequently involves nodal and extranodal sites. The presence of leukemic phase in ALCL is extremely rare and occurs exclusively with ALK1-positive ALCL. We describe two patients with ALK1-positive ALCL who developed a leukemic phase with rapid progression of the disease. Immunophenotypic pattern assessed on peripheral blood by flow cytometry revealed CD45, CD30, and CD25 positivity in both cases but NPM-ALK1 was expressed in only one case. Both patients developed leukemic phase as a terminal event of the disease and we share the immunophenotypic features of both cases.

Immunophenotypic profile of plasma cell leukemia: A retrospective study in a reference cancer center in India and review of literature.

Background: Plasma cell leukemia (PCL) is a rare but aggressive subtype of plasma cell dyscrasia. It is known to present with highly variable morphological features and may mimic with other lymphoid neoplasms. Multicolor flow cytometry (MFC) with availability of newer markers is highly useful in the diagnosis of the plasma cell leukemia. We present an immunophenotypic profile in ten cases of PCL along with their clinical and laboratory findings. Materials and Methods: We retrospectively studied immunophenotypic profile of 10 cases of plasma cell leukemia (out of 4615 cases of hematolymphoid neoplasms) using five parameter, three color flow cytometric analysis. We also studied their clinical presentation and other laboratory findings. Results: Common clinical features at presentation were weakness, bone pain, anemia, thrombocytopenia and osteolytic lesions. Plasma cell population was identified on strong expression of CD38 and co-expression of CD38 and CD138. CD56 was expressed in 44% cases. CD19 and CD20 were negative in all cases. Surface light chain restriction was seen in 50% cases and in remaining 50% cases revealed cytoplasmic light chain restriction. CD117 was expressed in one out of two cases studied. Conclusions: MFC immunophenotyping is highly useful to differentiate Plasma cell leukemia from other chronic lymphoproliferative disorders with plasmacytoid morphology as well as from non-neoplastic reactive PC and co-expression of CD38 and CD138 is a best combination to identify the plasma cells by MFC.

Immunophenotypic profile of plasma cell leukemia: A retrospective study in a reference cancer center in India and review of literature.

Background: Plasma cell leukemia (PCL) is a rare but aggressive subtype of plasma cell dyscrasia. It is known to present with highly variable morphological features and may mimic with other lymphoid neoplasms. Multicolor flow cytometry (MFC) with availability of newer markers is highly useful in the diagnosis of the plasma cell leukemia. We present an immunophenotypic profile in ten cases of PCL along with their clinical and laboratory findings. Materials and Methods: We retrospectively studied immunophenotypic profile of 10 cases of plasma cell leukemia (out of 4615 cases of hematolymphoid neoplasms) using five parameter, three color flow cytometric analysis. We also studied their clinical presentation and other laboratory findings. Results: Common clinical features at presentation were weakness, bone pain, anemia, thrombocytopenia and osteolytic lesions. Plasma cell population was identified on strong expression of CD38 and co-expression of CD38 and CD138. CD56 was expressed in 44% cases. CD19 and CD20 were negative in all cases. Surface light chain restriction was seen in 50% cases and in remaining 50% cases revealed cytoplasmic light chain restriction. CD117 was expressed in one out of two cases studied. Conclusions: MFC immunophenotyping is highly useful to differentiate Plasma cell leukemia from other chronic lymphoproliferative disorders with plasmacytoid morphology as well as from non-neoplastic reactive PC and co-expression of CD38 and CD138 is a best combination to identify the plasma cells by MFC.

Hypergranular precursor B-cell acute lymphoblastic leukemia in a 16-year-old boy.

Presence of cytoplasmic granules in the blasts is a well known feature of myeloid leukemia. ALL presenting with the numerous cytoplasmic granules in blasts is a rarity and may be misdiagnosed as acute myeloid leukemia. We describe a rare case of hypergranular precursor B-cell acute lymphoblastic leukemia (ALL) in an adolescent male expressing CD10, CD19, CytoCD22, CD34, as well as CD13 and CD117. The blasts were cytochemically negative for myeloperoxidase (MPO), and acid phosphatase (ACP) but were positive for non-specific esterase (NSE). In centers where immunophenotypic panel is usually decided on the basis of morphology with limited antibodies may result in an erroneous typing of such rare diseases. Hence it is important to be aware of this rare entity and to confirm the lineage of acute leukemia by using a comprehensive panel of antibodies for immunophenotypic analysis.

Intracytoplasmic antigen study by flow cytometry in hematolymphoid neoplasm.

Flow cytometric detection of intracellular antigens has become a standard method in establishing proper leukemic cell lineage affiliation. It has a non-debatable contribution to the diagnosis of hematolymphoid neoplasm as well as in minimal residual disease. Combination of analysis of fluorescence labeling and light scatter properties of cells allows rapid and better determination of target cell antigens. Regarding the detection of intracellular antigens, standardization of the procedure remains, however, a real challenge. Detection of intracellular antigens by flow cytometry (FCM) requires effective fixation and permeabilization of the cell membrane. In the available literature, some reports describe methodologies to achieve satisfactory results for detection of either cytoplasmic or nuclear antigens; however, no methodological consensus has yet been achieved among the laboratories. This article is an attempt to describe different approaches to detect intracellular molecules by FCM.

Exflagellated microgametes of Plasmodium vivax in human peripheral blood: a case report and review of the literature.

Peripheral blood smear examination is the most specific as well as the most common test performed for the diagnosis of malaria. Schizonts, ring forms (trophozoites) and gametocytes are the stages of malarial parasite that are commonly seen in the peripheral blood smear of a patient. Here, we report an extremely rare case of a 40-year-old male patient who presented with Plasmodium vivax infection with multiple exflagellated microgametes in the peripheral blood smear with review of the literature. Exflagellation of microgametes in malarial parasites is only seen in the definitive host, mosquito, and is very unusual to see during the developmental phases in the intermediate host, human. It is important to recognize these exflagellated microgametes in the peripheral blood smear as they may lead to diagnostic confusion with organisms such as spirochetes and trypanosomes.