<h3>Purpose</h3> The aim of our study is to assess the reliability of transcriptome profile platform outcomes between different tissue preservation conditions, RNA extraction methods and heart-failure-related genes, minimizing intra- and inter-experimental variability. <h3>Methods</h3> Our cohort of study is composed by 6 FFPE endomyocardial biopsies (EMBs) and 6 fresh frozen (FF) heart tissues samples belonging to the same adult end-stage heart failure patient, limiting inter-patient variability bias. Two major RNA extraction procedure were tested for FF and FFPE tissues. Then, we performed a high-throughput microarray expression profiling without a bias of transcript selection. We analyzed mRNAs expression through R programming and transcriptome analysis software, exploring >20000 transcripts. We refined the transcriptome analysis selecting, from a literature systematic review appraisal 30 heart-failure related genes, coupling the overlapping genes in the differential expression and network analysis. <h3>Results</h3> We compared the analysis of up- and down-regulated genes between the groups, addressing both the different RNA extraction procedure and tissue preservation, while sharing the same transcriptome platform. The differential expressed genes (DEG) analyzed had fold change set at 2 and -2, and a cut-off threshold of FDR adjusted p-value < 0,05. Heatmap showed a great samples' clusterization according to preservation and extraction methods. Only 284 genes passed filter criteria in FFPE inter-groups comparison of extraction methods, while 52 differed in FFPE vs. FF, showing a great overlap of transcriptome signature, corroborated by the heart-failure related set of genes. The network analysis highlighted that most of the DEG are strongly associated (p-value adjusted <0.004) to the dysregulation of reticulum and collagen related cell component, in accordance with heart failure stage. <h3>Conclusion</h3> To the best of our knowledge, this is the first study in heart transplantation fields comparing genetic extraction and preservation methods for microarray expression profile evaluation correlated with pathophysiological mechanism. A proof of concept that RNAs from FFPE-EMBs, and not only FF, are feasible for microarray expression profile evaluation, and derived transcriptome profiles have a strong potential for research and diagnostic clinical application.