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Filter-aided expansion proteomics

ABSTRACT: Hydrogel-based tissue expansion combined with mass spectrometry (MS) offers an emerging spatial proteomics approach. Here, we present a filter-aided expansion proteomics (FAXP) strategy for spatial proteomics analysis of archived formalin-fixed paraffin-embedded (FFPE) specimens. Compared to our previous ProteomEx method, FAXP employed a customized tip device to enhance both the stability and throughput of sample preparation, thus guaranteeing the reproducibility and robustness of the workflow. FAXP achieved a 14.5-fold increase in volumetric resolution. It generated over 8 times higher peptide yield and a 255% rise in protein identifications while reducing sample preparation time by 50%. We also demonstrated the applicability of FAXP using human colorectal FFPE tissue samples. Furthermore, for the first time, we achieved bona fide single-subcellular proteomics under image guidance by integrating FAXP with laser capture microdissection.

INSTRUMENT(S): timsTOF Pro

ORGANISM(S): Homo Sapiens (human) Mus Musculus (mouse)

TISSUE(S): Liver, Colon

DISEASE(S): Colon Cancer

SUBMITTER: Tiannan Guo

LAB HEAD: Tiannan Guo

PROVIDER: PXD041412 | Pride | 2024-10-01

REPOSITORIES: Pride

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			Action	DRS
	20230404_CRC_combined_protein_DDA.tsv	Tabular
	20230404_CRC_report.pg_matrix_DIA.tsv	Tabular
	20230404_Detection_range.zip	Other
	20230404_Detection_range_combined_protein_DDA.tsv	Tabular
	20230404_Detection_range_report.pg_matrix_DIA.tsv	Tabular

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Project description:Diethlstilbestrol (DES) is a synthetic estrogen prescribed to several millions of pregnant women worldwide. The risk for breast cancer after age 40 in women prenatally exposed to DES has been reported, however, the precise mechanism of susceptibilities to breast cancer remains to be resolved. To understand the mechanism of effect of intrauterine exposure to DES on human mammary glands at peripubertal stage, we investigated the global gene expression profile of terminal end buds (TEBs), the target of carcinogen, in rat mammary glamds at postnatal days (PND) 35 and 49, equivalent to peripubertal stage in human, exposed to low or high dose of DES at neonatal period. In all group, the number of TEBs was gradually increased and peaked at PND35 and decreased at PND49. At PND 35 and 49, low dose of DES-treated group showed highest number of TEBs. At PND35, β- and γ-casein mRNA expression inreased 8.19-fold and 26.05-fold, respectively, and the most significant network revealed by IPA analyses showed relevance of NF-κB in low dose of DES-treated group. The present findings suggested that deregulation of mammary gland differentiation and development-related genes group may contribute to the increase of number of TEBs at critical period of carcinogen susceptibilities. Experiment: We used laser capture microdissection to specifically isolate the cells from terminal end buds (TEBs) in rat mammary gland at 35 and 49 days after birth. Inbred Sprague-Dawley female rats were subcutaneously given 0.05ml sesame oil (group I, control) and either 1 μg (group II) or 100 μg (group III) diethlstilbestrol (DES, Sigma Chemical Co., St. Louis, Mo. USA) dissolved in 0.05 ml sesame oil within 24 hr of birth. Animal were weaned at 21 days of birth and each group of rats was autopsied at 35 and 49 days of birth. From group I (35 days of birth: n=2, 49 days of birth: n=2), group II (35 days of birth: n=2, 49 days of birth: n=2), group III (35 days of birth: n=2, 49 days of birth: n=2), we performed a hybridization, respectively.

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Filter-aided expansion proteomics

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