APPLICABILITY OF DIRECT IN-SITU REVERSE TRANSCRIPTION-POLYMERASE CHAIN-REACTION ON BONE-MARROW SMEARS

In situ reverse transcription (RT)-polymerase chain reaction (PCR) is a promising laboratory tool for biomedical investigation at the molecu lar level in tissues. Direct in-cell amplification of the breakpoint c luster region (BCR)-Abelson (ABL) fusion transcript of chronic myeloid leukemia (CML) has recently been accomplished in Italy using bone mar row mononuclear cell suspensions. The goals of this study are to deter mine if in situ RT-PCR amplification is possible on bone marrow spirat e smears and to demonstrate any unique factors in this procedure. A co mmercially available method was used because of the existence of publi shed protocols for adaptation. Bone marrow (BM) aspirate smears (n = 1 7) from patients with CML in blast crisis (positive case material) or other hematological malignancies (negative case material) were evaluat ed. Satisfactory amplification of the BCR-ABL fusion transcript occurr ed, and distinct blue cytoplasmic granules that varied in intensity we re found in most CML blasts. The negative case materials lacked the sp ecifically amplified granular signals. Overall signal strength and bac kgrounds were readily affected by the quality of the specimen as well as by changes in assay parameters. In conclusion, the direct in situ R T-PCR technique is applicable for bone marrow aspirate smear evaluatio n. However, it remains an investigative tool until optimization for se nsitivity, specificity, and accuracy can be achieved.