Common variable immunodeficiency associated enteropathy: A diagnostic enigma in developing countries.

Common variable immunodeficiency (CVID) is the most prevalent primary immunodeficiency disorder with different phenotypes and aetiologies. It is characterised by hypogammaglobulinaemia, defects in specific antibody response, erroneous activation and proliferation of T cells, leading to increased risk of recurrent infections. In CVID, "Variable" refers to the heterogeneity of clinical presentations, which include recurrent infections, autoimmunity, enteropathy, and increased risk of malignancies. This wide spectrum of disease manifestations and being a diagnosis of exclusion poses a diagnostic challenge. It is pertinent to mention that CVID along with associated complications is the commonest symptomatic primary antibody deficiency but is scarcely mentioned in local literature. The main aim of presenting this case is to impress upon the importance of systematic immunological workup in cases of suspected immunodeficiency to prevent morbidity and mortality.

Frequency and Aberrant CD Marker Profile of Acute Leukaemias.

OBJECTIVE: To determine the frequency of different types of acute leukaemia and their subtypes along with associated aberrant CD markers. STUDY DESIGN: Descriptive study. Place and Duration of the Study: Department of Immunology Armed Forces Institute of Pathology, National University of Medical Sciences, Rawalpindi, Pakistan, from November 2021 to October 2023. METHODOLOGY: All samples received for flow cytometric immunophenotyping with suspicion of acute leukaemia were included in the study. Cells were stained with fluorochrome labelled monoclonal antibodies against lineage-specific cluster of differentiation (CD) markers through a lyse-wash procedure. Acquisition and analysis were done using multi-parameter BD FACS Canto II Flow cytometer and BD FACS Diva software, respectively. Data were entered and analysed using SPSS v 23.0. RESULTS: Over a period of 2 years, a total of 1,115 suspected patients were tested for acute leukaemia. Among them, 728 (65.3%) were males and 387 (34.7%) were females, with mean age 28 ± 21 years, ranging from 1 week to 87 years. Among a total of 875/1115 (78.5%) diagnosed cases of acute leukaemia, AML was the most common leukaemia present in 408/875 (46.6%) patients followed by B-ALL and T-ALL in 384/875 (43.8%) and 70/87 (8%) patients, respectively (p = 0.5712). Aberrant CD markers were detected in 109/875 (12.5%) leukaemias (p = 0.0628). The most common aberrant CD markers in B-ALL were CD13 and CD33 present in 30/384 (7.8%) cases separately. Among AML and T-ALL most common aberrant CD markers were CD7 and CD33 present in 25/408 (6.13%) and 7/70 (10%) cases, respectively. CONCLUSION: Special consideration should be given to the presence of aberrant CD markers when assigning lineages to acute leukaemias. They may be important diagnostic, prognostic, and management tools for institution of immunotherapy. KEY WORDS: Aberrant CD markers, Acute leukaemia, CD Markers, Flow cytometry, Immunophenotyping.

Development and Validation of In-house HEp-2 Cell Slides for Detection of Antinuclear Antibodies (ANA) by Indirect Immunofluorescence.

OBJECTIVE: To develop and validate in-house HEp-2 cell slides for the detection of ANA by indirect immunofluorescence. STUDY DESIGN: Cross sectional validation study. Place and Duration of the Study: Department of Immunology, Armed Forces Institute of Pathology Rawalpindi, Punjab, Pakistan, from April to September 2022. METHODOLOGY: This study involved development of in-house HEp-2 cell slides after procuring cell lines, sub-culturing and fixing them on different slides using variety of fixatives under different protocols. After standardisation of procedure, validation of procedure was done by testing sera of 305 patients for ANA detection at 1:40 dilution on in-house HEp-2 cell slides and subsequently on commercial HEp-2 cell slides (gold standard). Indirect immunofluorescence was observed by the two observers working independently and kept blinded from the results interpreted by each other. Data were collected on a pre-designed proforma and then analysed. Sensitivity, specificity, and positive predictive value (PPV) and negative predictive values (NPV) of in-house HEp-2 cell slides were calculated. RESULT: Sera of 305 patients were tested on in-house and commercial HEp-2 cell slides. Sensitivity and specificity of in-house HEp-2 cell slides for ANA detection were 96.92% and 99.58%, respectively. PPV and NPV of in-house HEp-2 cell slides came out to be 98.43% and 99.17% respectively. CONCLUSION: In-house HEp-2 cell slides are as effective as commercial HEp-2 cell slides for the detection of ANA and can be used as cost-effective alternative. KEY WORDS: Antinuclear antibodies (ANA), Human epithelial type-2 (HEp-2), Immunofluorescence.

T cell reactivity to Collagen II as a possible prognostic marker in patients with rheumatoid arthritis.

OBJECTIVE: To compare the frequency and the functional state of the collagen II reactive T cells with disease activity in rheumatoid arthritis patients and healthy controls. METHODS: This case-control cross-sectional study was carried out at the Department of Immunology; Armed Forces Institute of Pathology, Rawalpindi, Pakistan, from June to October 2014. Rheumatologist from Rehmat Noor Rheumatology Clinic, a private health facility of the city, was requested to send in patients with clinical diagnosis of rheumatoid arthritis. Samples were obtained and relevant investigations were carried out. Data were compared with a group of age and gender-matched healthy subjects. T cell proliferative response was assessed against bovine collagen II by measuring incorporation of bromodeoxyuridine into deoxyribonucleic acid of proliferating cells and by expression of CD25 on proliferating cells as percentage of CD3+/bromodeoxyuridine+ and CD3+/CD25+ T-cells, respectively. Among the patients, the frequency of T cells with disease activity was compared. Patients were classified into groups of mild, moderate and severe disease and frequency of CD3+/bromodeoxyuridine+, frequency of CD3+/CD25+ cells, mean fluorescent intensity of bromodeoxyuridine-fluorescein isothiocyanate and mean fluorescent intensity of CD25-fluorescein isothiocyanate were compared in the groups. RESULTS: Of the 60 subjects, 30(50%)were patients and 30(50%) were controls. Of the patients, 5(16.66%) were males and 25(83.33%) were females with an overall mean age of 42±12 years. The mean age of the controls was 41±9.28 years. Mean disease duration of the patients was 10.5 ± 4.2 years. Percentage of CD3+/CD25+ cells and CD3+/bromodeoxyuridine+ cells stimulated with collagen II, in patients was much higher than the controls(p<0.05).Statistically significant differences were observed when frequency of CD3+/bromodeoxyuridine+ cells and CD3+/CD25+ cells was compared among the mild, moderate and severe patient groups (p<0.05). CONCLUSIONS: Collagen II was found to be an important auto antigen in joints of rheumatoid arthritis patients.