title: Histology-guided protein digestion/extraction from formalin-fixed and paraffin-embedded pressure ulcer biopsies.
abstract: Herein we present a simple, reproducible and versatile approach for in situ protein digestion and identification on formalin-fixed and paraffin-embedded (FFPE) tissues. This adaptation is based on the use of an enzyme delivery platform (hydrogel discs) that can be positioned on the surface of a tissue section. By simultaneous deposition of multiple hydrogels over select regions of interest within the same tissue section, multiple peptide extracts can be obtained from discrete histological areas. After enzymatic digestion, the hydrogel extracts are submitted for LC-MS/MS analysis followed by database inquiry for protein identification. Further, imaging mass spectrometry (IMS) is used to reveal the spatial distribution of the identified peptides within a serial tissue section. Optimization was achieved using cutaneous tissue from surgically excised pressure ulcers that were subdivided into two prime regions of interest: the wound bed and the adjacent dermal area. The robust display of tryptic peptides within these spectral analyses of histologically defined tissue regions suggests that LC-MS/MS in combination with IMS can serve as useful exploratory tools.
authors.affiliation: Dipartimento di Chimica e Tecnologie Chimiche, Università della Calabria, Arcavacata di Rende, Italy.|Mass Spectrometry Research Center, Vanderbilt School of Medicine, Nashville, TN, USA.
authors.forename: Domenico
authors.initials: D
authors.lastname: Taverna
authors.affiliation: Department of Plastic Surgery, Vanderbilt School of Medicine, Nashville, TN, USA.
authors.forename: Alonda C
authors.initials: AC
authors.lastname: Pollins
authors.affiliation: Department of Plastic Surgery, Vanderbilt School of Medicine, Nashville, TN, USA.|Department of Cell & Developmental Biology, Vanderbilt School of Medicine, Nashville, TN, USA.
authors.forename: Lillian B
authors.initials: LB
authors.lastname: Nanney
authors.affiliation: Dipartimento di Chimica e Tecnologie Chimiche, Università della Calabria, Arcavacata di Rende, Italy.
authors.forename: Giovanni
authors.initials: G
authors.lastname: Sindona
authors.affiliation: Mass Spectrometry Research Center, Vanderbilt School of Medicine, Nashville, TN, USA.|Department of Biochemistry, Vanderbilt School of Medicine, Nashville, TN, USA.
authors.forename: Richard M
authors.initials: RM
authors.lastname: Caprioli
mesh_terms: D001706:Biopsy / Q000379:methods*; D002853:Chromatography, Liquid; D004801:Eosine Yellowish-(YS); D005557:Formaldehyde; D006416:Hematoxylin; D006801:Humans; D020100:Hydrogels; D016612:Paraffin Embedding; D003668:Pressure Ulcer / Q000378:metabolism* / Q000473:pathology; D011506:Proteins / Q000302:isolation & purification*; D040901:Proteomics / Q000379:methods*; D013048:Specimen Handling; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013194:Staining and Labeling; D053719:Tandem Mass Spectrometry; D016707:Tissue Fixation; D014357:Trypsin / Q000494:pharmacology
chemical_list: D020100:Hydrogels; D011506:Proteins; D005557:Formaldehyde; D014357:Trypsin; D004801:Eosine Yellowish-(YS); D006416:Hematoxylin
keywords: chronic wounds; hydrogel; imaging mass spectrometry; pressure ulcers; proteomics