A non-hazardous deparaffinization protocol enables quantita-tive proteomics of core needle biopsy-sized formalin-fixed and paraffin embedded (FFPE) tissue specimens.

ABSTRACT: Most human tumor tissues that are obtained for pathology and diagnostic purposes are forma-lin-fixed and paraffin embedded (FFPE). To perform quantitative proteomics of FFPE samples, paraffin has to be removed and formalin-induced crosslinks have to be reversed prior to prote-olytic digestion. A central component of almost all deparaffinization protocols is xylene, a toxic and highly flammable solvent that has been reported to negatively affect protein extraction and quantitative proteome analysis. Here, we present a ‘green’ xylene-free protocol for accel-erated sample preparation of FFPE tissues based on paraffin-removal with hot water. Com-bined with tissue homogenization using disposable micropestles and a modified protein aggre-gation capture (PAC) digestion protocol, our workflow enables streamlined and reproducible quantitative proteomic profiling of FFPE tissue. Label free quantitation of FFPE cores from human ductal breast carcinoma in-situ (DCIS) xenografts with a volume of only 0.79 mm3 showed a high correlation between replicates (r2=0.992) with a median %CV of 16.9%. Im-portantly, this small volume is amenable to tissue micro array (TMA) cores and core needle bi-opsies, while our results and the easy-of-use indicate that a further downsizing is feasible. Fi-nally, our FFPE workflow does not require costly equipment and can be established in every standard clinical laboratory.

INSTRUMENT(S): Q Exactive Plus

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Mammary Ductal Carcinoma Cell

DISEASE(S): Breast Cancer

SUBMITTER: Georgia Mitsa  

LAB HEAD: Christoph H Borchers

PROVIDER: PXD031946 | Pride | 2022-05-19

REPOSITORIES: Pride