Detection of Central Nervous System Leukemia in Children with Acute Lymphoblastic Leukemia by Real-Time Polymerase Chain Reaction Sharon R. Pine, Changhong.

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Detection of Central Nervous System Leukemia in Children with Acute Lymphoblastic Leukemia by Real-Time Polymerase Chain Reaction Sharon R. Pine, Changhong Yin, Yousif H. Matloub, Hatem E. Sabaawy, Claudio Sandoval, Oya Levendoglu-Tugal, M. Fevzi Ozkaynak, Somasundaram Jayabose The Journal of Molecular Diagnostics Volume 7, Issue 1, Pages 127-132 (February 2005) DOI: 10.1016/S1525-1578(10)60018-9 Copyright © 2005 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 1 Real-time PCR assays for determination of the most suitable storage condition for DNA stability in CSF. Samples were amplified using primers specific to the human β-globin gene. Amplification plot of cycle number versus fluorescence shows PCR results of dilutions of standard human genomic DNA (Roche) down to 10 starting template copies and DNA extracted from equal aliquots of CSF stored in 1:1 RPMI (for 2 days and 7 days with shipping) or with no medium (for 1 hour and 7 days) (A), or CSF stored in 25% ethanol for 2 days without shipping, and 2 and 7 days with shipping (B). Standards were amplified in duplicates and samples were amplified in triplicates but results are shown in singles for simplification. The cycle number in which the fluorescence level surpasses the crossing line is inversely proportional to the number of PCR DNA targets at the beginning of the reaction. NP, no preservatives; NBM, normal bone marrow; EOI, end of induction. The Journal of Molecular Diagnostics 2005 7, 127-132DOI: (10.1016/S1525-1578(10)60018-9) Copyright © 2005 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

Figure 2 Quantitative real-time PCR analysis of CNS-leukemia involvement in pediatric ALL. Amplification plot of cycle number versus fluorescence shows real-time PCR results for patient-specific amplification of the Vδ2-Dδ2 clonal rearrangement identified at diagnosis in patient 28. Curves represent dilutions of patient diagnosis bone marrow DNA down to 1 copy PCR targets (diluted in a background of 20,000 genomic copies of non-leukemic DNA), patient diagnosis and end of induction CSF bone marrow DNA samples, and a normal bone marrow negative control DNA. Standards and patient samples were amplified in triplicates, but standards are shown in duplicates and samples are shown in singles, for simplification. The cycle number in which the fluorescence level surpasses the crossing line is inversely proportional to the number of PCR DNA targets at the beginning of the reaction. EOI, end of induction; NBM, normal bone marrow. The Journal of Molecular Diagnostics 2005 7, 127-132DOI: (10.1016/S1525-1578(10)60018-9) Copyright © 2005 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions