Comparative analysis of fluorescence in situ hybridization and real time polymerase chain reaction in diagnosis of chronic myeloid leukemia

Objective: To compare the sensitivity and specificity of fluorescence in situ hybridization [FISH] with real time polymerase chain reaction [RT-PCR] in the diagnosis of Chronic Myeloid Leukemia [CML]

Study Design: A cross-sectional, analytical study

Place and Duration of Study: Haematology Department, Armed Forces Institute of Pathology, Rawalpindi, from January 2012 to February 2014

Methodology: A total number of 87 patients of CML were studied. The diagnosis was made on the basis of clinical history, peripheral blood and bone marrow aspiration. These patients were tested for the presence of BCR-ABL1 fusion gene by RT-PCR and FISH. About 5 ml of venous blood was collected, half was taken in heparin for FISH and half in ethylenediamine tetra-acetic acid [EDTA] for CBC and PCR. For FISH, cells were cultured for 24 hours in RPMI 1640 medium and evaluated using BX51 fluorescence microscope for dual fusion signal of yellow colour. Samples having 20 or more interphases positive for dual fusion signals were taken as positive. For PCR, RNA extraction was done by Tri-Reagent LS [MRC, USA] and cDNA was synthesized using reverse transcriptase and gene specific primer. RT-PCR was done on ABI-7500. The positive samples were identified when fluorescence exceeded threshold limit. Results of RT-PCR and FISH were compared

Results: Out of the 87 patients, 85 [97.7%] were PCR positive and 2 [2.3%] were PCR negative, whereas in FISH 83 [95.4%] were positive and 4 [4.5%] were negative. Sensitivity and specificity of FISH was 97.6% and 100%, respectively

Conclusion: FISH is a reliable supplementary method to PCR for detection of BCR-ABL1 fusion gene in the diagnosis of CML

Early diagnosis of dengue virus infection in clinically suspected cases

Objective: Comparison of real time reverse transcriptase polymerase chain reaction [RTPCR] and immunoglobulin M [IgM] capture enzyme linked immunosorbent assay [ELISA] for diagnosis of dengue virus infection in first week of illness in clinically suspected patients of dengue fever

Study Design: Cross sectional study

Place and Duration of Study: Department of haematology, Armed Forces Institute of Pathology [AFIP] Rawalpindi from Jan 2013 to Nov 2013

Material and Methods: A cross sectional study including 68 clinically suspected patients of dengue fever according to the World Health Organization [WHO] criteria. IgM capture ELISA and RT PCR for dengue virus ribonucleic acid [RNA] was performed on samples collected from patients having fever for 1 to 7 days. These were divided into two groups. Patients in group 1 presented with fever of 4 days or less, patients in group 2 had fever of 5 to 7 days duration

Results: In group 1, 72% of the patients were positive by RT PCR while 31% were positive by IgM capture ELISA. In group 2, 43% of the patients were positive by RT PCR while 97% were positive by ELISA

Conclusion: RT PCR can be used for early detection of dengue virus infection in the first few days of fever while IgM ELISA is diagnostic afterwards