Gene expression profiling is a robust technology for the diagnosis of hematologic malignancies with high accuracy. It may complement current diagnostic algorithms and could offer a reliable platform for patients who lack access to today's state-of-the-art diagnostic work-up. Our comprehensive gene expression data set will be submitted to the public domain to foster research focusing on the molecular understanding of leukemias.
SummaryGene expression profiling has the potential to enhance current methods for the diagnosis of haematological malignancies. Here, we present data on 204 analyses from an international standardization programme that was conducted in 11 laboratories as a prephase to the Microarray Innovations in LEukemia (MILE) study. Each laboratory prepared two cell line samples, together with three replicate leukaemia patient lysates in two distinct stages: (i) a 5-d course of protocol training, and (ii) independent proficiency testing. Unsupervised, supervised, and r 2 correlation analyses demonstrated that microarray analysis can be performed with remarkably high intra-laboratory reproducibility and with comparable quality and reliability.
The diagnosis of myelodysplastic syndrome (MDS) currently relies primarily on the morphologic assessment of the patient's bone marrow and peripheral blood cells. Moreover, prognostic scoring systems rely on observer-dependent assessments of blast percentage and dysplasia. Gene expression profiling could enhance current diagnostic and prognostic systems by providing a set of standardized, objective gene signatures. Within the Microarray Innovations in LEukemia study, a diagnostic classification model was investigated to distinguish the distinct subclasses of pediatric and adult leukemia, as well as MDS. Overall, the accuracy of the diagnostic classification model for subtyping leukemia was approximately 93%, but this was not reflected for the MDS samples giving only approximately 50% accuracy. Discordant samples of MDS were classified either into acute myeloid leukemia (AML) or "none-of-thetargets" (neither leukemia nor MDS) categories. To clarify the discordant results, all submitted 174 MDS samples were externally reviewed, although this did not improve the molecular classification results. However, a significant correlation was noted between the AML and "none-ofthe-targets" categories and prognosis, leading to a prognostic classification model to predict for time-dependent probability of leukemic transformation. The prognostic classification model accurately discriminated patients with a rapid transformation to AML within 18 months from those with more indolent disease.
Summary. Radioimmunoassays for erythropoietin are limited so far to a few specialized laboratories and this requires transport and storage of samples. We therefore tested the stability of immunoreactive erythropoietin in serum and plasma samples obtained from a uremic and a nonuremic anemic patient. No significant change in the concentration of immunoreactive erythropoietin was found in either serum or plasma samples for up to 14 days of storage. This type of stability was observed no matter whether the samples were stored at room temperature, 4 ° C, or -20 ° C. There was no difference between the estimates of erythropoietin in serum and heparinized plasma. Validity of the radioimmunoassay used in this study was demonstrated by parallelism of dilution curves of test specimens and the 2 na International Reference Preparation for erythropoietin and by a close correlation between the immunoreactivity and the bioactivity of the hormone, as assessed in the same samples by the exhypoxic polycythemic mouse bioassay.In conclusion the data obtained clearly indicate that the necessity of storage and transport of clinical samples does not limit the practicability of the radioimmunoassay for erythropoietin.Key words: Erythropoietin -Stability -Radioimmunoassay -Polycythemic mouse bioassay -Recombinant DNA Since recombinant human erythropoietin (rhEPO) became available for replacement therapy in paAbbreviations. BSA = bovine serum, albumin; EPO = erythropoietin; irEPO=immunoreactive erythropoietin; IRP=Inter-national Reference Preparation; hct=hematocrit; rhEPO=re-combinant human erythropoietin; RIA = radioimmunoassay tients with certain forms of anemia [5,9], the determination of erythropoietin (EPO) levels in body fluids has gained increasing clinical importance. However, radioimmunoassay kits are not yet commercially available and the limitation of EPO determinations to specialized laboratories necessitates storage and transport of the samples. This makes it important to know the stability of EPO in human samples. We therefore performed subsequent radioimmunological determinations of EPO for up to 14 days on serum and plasma samples which were stored at room temperature, 4 ° C, or -20 ° C. The validity of RIA estimates was secured by determination of the biological activity of the same samples using an in vivo bioassay and by comparison of the slopes of dilution curves of test samples and the 2 nd International Reference Preparation (IRP) for EPO.
Material and Methods
Antiserum to Erythropoietin
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