360 Waste not, test more: Innovations in tissue processing to expand the testing of clinical specimens

Abstract

OBJECTIVES/GOALS: Clinical tissue specimens are primarily destined for formalin fixed, paraffin embedded processing to create a basis for diagnosis by microscopic examination. Innovations in specimen processing are required to expand its availability for inclusion as the substrate in assays that can contribute to the further development of Precision Medicine. METHODS/STUDY POPULATION: Transurethral resection of bladder tumors were selected for testing based on availability and tissue composition. A wash step was used to generate daughter aliquots composed of dislodged cells and a solution with prior contact to the parent tissue. This wash step served two purposes: 1) reduce the amount of contaminating material from spreading to other cases, a problem known to be associated with this type of specimen; and 2) create aliquots from which additional informative data could be generated. These daughter aliquots were then examined to determine their value as a source for exosome profiling, metabolomic studies, molecular characterization and organoid development. The parent tissue was not compromised, was able to undergo conventional processing and yielded results equivalent to unwashed specimens. RESULTS/ANTICIPATED RESULTS: Exosomes secreted by the tumor cells were identified to be present in the daughter aliquots by a combination of their isolation using CD31 and detection of miR-21 expression. These exosomes were confirmed to be not related to fragmented cells from testing for beta-tubulin. A global/discovery-based approach using mass spectrometry provided insights into early characterization of metabolomic profiles present in these tumor cells. Ample amounts of high quality DNA (226 ng/ul concentrations; 11.3 ug total) were recovered from the dislodged, excess cells in the wash for molecular studies. Finally, from viable cells recovered in one of the daughter wash aliquots, the ability to grow organoids was proven to be possible and reproducible. DISCUSSION/SIGNIFICANCE: Based on these results, the value of the clinical specimen can be markedly expanded for utilization in research and possible clinical use without detracting from the parent tissue. This non-destructive, easy to adopt wash procedure can potentially lead to an influx of data that may ultimately prove useful in improving patient care.