Achieving the best RNA quality in urologic tumor samples intended for transcriptome analysis

Abstract

Purpose: To conduct research on the molecular oncology, physiology, and immunology of urologic tumors requires dissociated viable samples. Improper collection compromises the quality of data attained in molecular and functional assays due to the increased quantities of degraded proteins and RNA. We sought to improve the methods for tissue collection which can avoid generating considerable loss in the viability of cells for further analyses. Materials and Methods: Fifty resected tumor samples from 35 patients were obtained with different surgical techniques and at various time points for viability and RNA quality evaluation. The degradation of RNA was evaluated by its Qubit IQ score, OD 260/280 ratio, total yield, and quantity of β-actin. Results: Snap-frozen tissue samples obtained within 30 min showed better cell viability (P < 0.0001), RNA total yield (P = 0.0081), Qubit ratio (P = 0.003), OD 260/280 ratio (P = 0.4213), and quantity of β-actin (P = 0.0015). Moreover, the bladder tumor samples collected from transurethral biopsy presented more satisfied cell viability results than the ones resected by transurethral electroresection (P < 0.0001). Conclusion: Tumor samples should be processed or frozen freshly within 30 min once removed from human body. Furthermore, transurethral biopsy of bladder tumor is considered a better method for collecting samples for further molecular oncology studies. The high-quality RNA produced enable researchers to conduct more reliable studies by avoiding the experimental artifacts due to the presence of cellular debris or dead cells.