Figure 4.
Figure 4. Analysis of EHZF expression in a blood disease profiling array. (A) A radiolabeled probe for EHZF was hybridized to a blood disease profiling array (cat. no. #7842-1; BD Biosciences). The filter was then washed and exposed for 48 hours. Diseases represented on the array are acute myelogenous leukemia (AML), chronic myelogenous leukemia (CML), Hodgkin disease (HD), non-Hodgkin lymphoma (NHL), and von Willebrand disease (VWD). Each sample includes cDNAs from purified monocytes (CD14+), B and T lymphocytes (CD19+ and CD3+, respectively), mononuclear cells (MNs), polymorphonucleates (PNs), and total leukocytes (TLs). Lymph node (LN) samples were from healthy subjects and from HD02, HD04, HD06, HD09, and NHL02 patient samples only. The filter was then later stripped in 0.5% SDS by boiling and the hybridized with a random primer-labeled probe for EBF, which bound only to the cDNAs from CD19+ B lymphocytes. Subsequent stripping and hybridization of the filter with ubiquitin probe confirmed the uniform presence of cDNA in the filter spots. (B) RT-PCR analysis of EHZF expression in acute myelogenous leukemia (AML). cDNA was amplified by PCR in duplicate from AML RNA samples with primers specific for EHZF for 40 cycles or with GAPDH primers for 24 cycles. PCR products were analyzed by 1% agarose gels and ethidium bromide staining. As controls, 1 ng of plasmids pFlag-CS2-OAZ(KIAA0760-L), p3xFlagCMV7.1-EHZF, or buffer containing no plasmid (control) were amplified for 20 cycles.

Analysis of EHZF expression in a blood disease profiling array. (A) A radiolabeled probe for EHZF was hybridized to a blood disease profiling array (cat. no. #7842-1; BD Biosciences). The filter was then washed and exposed for 48 hours. Diseases represented on the array are acute myelogenous leukemia (AML), chronic myelogenous leukemia (CML), Hodgkin disease (HD), non-Hodgkin lymphoma (NHL), and von Willebrand disease (VWD). Each sample includes cDNAs from purified monocytes (CD14+), B and T lymphocytes (CD19+ and CD3+, respectively), mononuclear cells (MNs), polymorphonucleates (PNs), and total leukocytes (TLs). Lymph node (LN) samples were from healthy subjects and from HD02, HD04, HD06, HD09, and NHL02 patient samples only. The filter was then later stripped in 0.5% SDS by boiling and the hybridized with a random primer-labeled probe for EBF, which bound only to the cDNAs from CD19+ B lymphocytes. Subsequent stripping and hybridization of the filter with ubiquitin probe confirmed the uniform presence of cDNA in the filter spots. (B) RT-PCR analysis of EHZF expression in acute myelogenous leukemia (AML). cDNA was amplified by PCR in duplicate from AML RNA samples with primers specific for EHZF for 40 cycles or with GAPDH primers for 24 cycles. PCR products were analyzed by 1% agarose gels and ethidium bromide staining. As controls, 1 ng of plasmids pFlag-CS2-OAZ(KIAA0760-L), p3xFlagCMV7.1-EHZF, or buffer containing no plasmid (control) were amplified for 20 cycles.

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