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The rapid diagnosis of acute promyelocytic leukaemia using PML (5E10) monoclonal antibody
S J O'Connor1, P D Forsyth, S Dalal
1Department of Haematology, The General Infirmary at Leeds.
Abstract:
Acute promyelocytic leukaemia (APL) is characterized cytogenetically by t(15;17)(q22:q21) which results in the production of a PML/RAR alpha fusion protein. Detection of the translocation or the fusion gene product is required for objective diagnosis of APL. This can be accomplished by conventional cytogenetic methods, fluorescence in situ hybridization or RT-PCR. Such techniques are time consuming and not universally available. The intracellular distribution of the PML protein in promyelocytes is characteristically altered in APL and this can be detected by immunocytochemistry. We have assessed two immunocytochemical methods, immunofluorescence and alkaline phosphatase-anti-alkaline phosphatase staining (APAAP), with regard to sensitivity, specificity and rapidity of diagnosis. 85 patients with AML including 15 cases of APL were studied. Immunofluorescence PML detection was concordant with RT-PCR for t(15:17) in 14/15 (93.3%) cases with no false positives. The negative APL case in our series was a patient with a 5' PML breakpoint who did not express the reciprocal t(17;15) fusion product. APAAP was concordant in only 6/13 (46%) APL cases with one false positive. In conclusion, immunofluorescent localization of PML using 5E10 monoclonal antibody is a rapid, sensitive and specific diagnostic tool for APL.
Insights
Immunofluorescence detection of PML protein is a rapid, sensitive, and specific diagnostic tool for acute promyelocytic leukemia (APL). This method offers a faster alternative to traditional techniques for diagnosing APL, a cancer characterized by a specific genetic translocation.
Area of Science:
- Hematology
- Molecular Biology
- Oncology
Background:
- Acute promyelocytic leukemia (APL) is defined by the t(15;17) chromosomal translocation, leading to a PML/RAR alpha fusion protein.
- Accurate diagnosis of APL requires detecting this translocation or fusion protein, often through time-consuming methods like cytogenetics, FISH, or RT-PCR.
- Alterations in the intracellular distribution of PML protein in promyelocytes are characteristic of APL and can be assessed via immunocytochemistry.
Purpose of the Study:
- To evaluate the sensitivity, specificity, and rapidity of two immunocytochemical methods for diagnosing APL.
- To compare immunofluorescence and APAAP staining for PML protein detection against established molecular diagnostic techniques.
Main Methods:
- Assessed immunofluorescence and alkaline phosphatase-anti-alkaline phosphatase (APAAP) staining for PML protein.
- Studied 85 acute myeloid leukemia (AML) patients, including 15 with APL.
- Correlated immunocytochemical results with RT-PCR detection of the t(15;17) translocation.
Main Results:
- Immunofluorescence demonstrated 93.3% concordance with RT-PCR for t(15;17) in APL cases, with no false positives.
- One APL case negative by immunofluorescence had a 5' PML breakpoint, lacking the reciprocal fusion product.
- APAAP staining showed only 46% concordance in APL cases, with one false positive.
Conclusions:
- Immunofluorescent localization of PML protein using the 5E10 monoclonal antibody is a rapid, sensitive, and specific diagnostic tool for APL.
- This immunocytochemical approach offers a valuable alternative to conventional diagnostic methods for APL.
- The findings highlight the utility of immunofluorescence in the timely and accurate diagnosis of APL.