BACKGROUND: Gene expression profiling is a useful tool for cancer diagnosis and basic research. A major limitation is that, even during short-term storage of native specimens of peripheral blood or bone marrow (BM) and/or RNA isolation, significant changes of gene expression pattern can occur because of gene induction, repression, and RNA degradation. METHODS: We investigated the effectiveness of a newly developed RNA stabilization and preparation system for BM specimens (PAXgene Bone Marrow RNA System) over time. We analyzed 256 RNA samples, processed from 64 BM specimens. RESULTS: Although the overall RNA yield (normalized to 1 x 10(7) leukocytes) was not different, the RNA preparation using unstabilized reference samples had an approximately 3 times higher failure rate. With the PAXgene system, we observed significantly higher RNA integrity compared with the reference RNA preparation system (P <0.01). In the stabilized samples, we found very high pairwise correlation in gene expression (DeltaDeltaC(T) 0.16-0.53) for the analyzed genes (GATA1, RUNX1, NCAM1, and SPI1) after 48-h storage compared with immediate preparation of RNA (2 h after BM collection). However, we found major differences in half of the analyzed genes using the reference RNA isolation procedure (DeltaDeltaC(T) 1.07 and 1.32). CONCLUSIONS: The PAXgene system is able to stabilize RNA from clinical BM samples and is suitable to isolate high-quality and -quantity RNA.
BACKGROUND: Gene expression profiling is a useful tool for cancer diagnosis and basic research. A major limitation is that, even during short-term storage of native specimens of peripheral blood or bone marrow (BM) and/or RNA isolation, significant changes of gene expression pattern can occur because of gene induction, repression, and RNA degradation. METHODS: We investigated the effectiveness of a newly developed RNA stabilization and preparation system for BM specimens (PAXgene Bone Marrow RNA System) over time. We analyzed 256 RNA samples, processed from 64 BM specimens. RESULTS: Although the overall RNA yield (normalized to 1 x 10(7) leukocytes) was not different, the RNA preparation using unstabilized reference samples had an approximately 3 times higher failure rate. With the PAXgene system, we observed significantly higher RNA integrity compared with the reference RNA preparation system (P <0.01). In the stabilized samples, we found very high pairwise correlation in gene expression (DeltaDeltaC(T) 0.16-0.53) for the analyzed genes (GATA1, RUNX1, NCAM1, and SPI1) after 48-h storage compared with immediate preparation of RNA (2 h after BM collection). However, we found major differences in half of the analyzed genes using the reference RNA isolation procedure (DeltaDeltaC(T) 1.07 and 1.32). CONCLUSIONS: The PAXgene system is able to stabilize RNA from clinical BM samples and is suitable to isolate high-quality and -quantity RNA.
Authors: Helen M Moore; Andrea B Kelly; Scott D Jewell; Lisa M McShane; Douglas P Clark; Renata Greenspan; Daniel F Hayes; Pierre Hainaut; Paula Kim; Elizabeth Mansfield; Olga Potapova; Peter Riegman; Yaffa Rubinstein; Edward Seijo; Stella Somiari; Peter Watson; Heinz-Ulrich Weier; Claire Zhu; Jim Vaught Journal: J Proteome Res Date: 2011-06-21 Impact factor: 4.466
Authors: Helen M Moore; Andrea Kelly; Scott D Jewell; Lisa M McShane; Douglas P Clark; Renata Greenspan; Pierre Hainaut; Daniel F Hayes; Paula Kim; Elizabeth Mansfield; Olga Potapova; Peter Riegman; Yaffa Rubinstein; Edward Seijo; Stella Somiari; Peter Watson; Heinz-Ulrich Weier; Claire Zhu; Jim Vaught Journal: Biopreserv Biobank Date: 2011-04 Impact factor: 2.300