proteome profiler human protease array kit  (R&D Systems)

Journal: iScience

Article Title: Human monocyte-derived macrophages shift subcellular metalloprotease activity depending on their activation state

doi: 10.1016/j.isci.2024.111171

Figure Lengend Snippet: Proteases and protease inhibitors/activators are increased in the cell culture supernatants and cell lysate of M(IL-4) compared to M(LPS/IFN-γ) (A) Protease and protease regulators detected in cell culture supernatants of M(M-CSF), M(LPS/IFN-γ), and M(IL-4) by proteome profiler array analysis. Expression was measured in comparison to the internal array controls and depicted as a scale bar from 0% to 100%. White squares indicate undetectable expression levels. (B) Quantification of secreted proteases and regulators in cell culture supernatants of M(M-CSF), M(LPS/IFN-γ), and M(IL-4) by gelatin in gel zymography (for MMP-9 protein levels) and ELISA (for MMP-12, cathepsin D, TIMP-1, TIMP-2, serpin E1, cystatin C). Each data point represents cells from a different donor (at least n = 7). Error bars are mean ± SEM. p values less than 0.05 were considered significant (∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001; ns non-significant). (C) Quantification of secreted MMP-9/TIMP-1 complex levels in cell culture supernatants of M(M-CSF), M(LPS/IFN-γ), and M(IL-4) by ELISA. Each data point represents cells from a different donor ( n = 9). Error bars are mean ± SEM. p values less than 0.05 were considered significant (∗ p < 0.05; ns non-significant). (D) Analysis of proteases and protease regulators in cell lysates of M(M-CSF), M(LPS/IFN-γ), and M(IL-4), measured using proteome profiler arrays. Expression was measured in comparison to the internal array controls and depicted as a scale bar from 0% to 100%. White squares indicate undetectable expression levels. (E) Quantification of cell-associated proteases and regulators of M(M-CSF), M(LPS/IFN-γ), and M(IL-4) by gelatin in gel zymography (for MMP-9 protein levels), ELISA (for MMP-12, ADAM-9, TIMP-2, serpin E1, and cystatin C) and flow cytometry (for EMMPRIN). Each data point represents cells from a different donor (at least n = 8). Error bars are mean ± SEM. p values less than 0.05 were considered significant (∗ p < 0.05; ∗∗ p < 0.01; ∗∗∗ p < 0.001; ∗∗∗∗ p < 0.0001; ns non-significant). (F) MMP-9 protein quantification by gelatin in gel zymography in the cell lysate of M(M-CSF), M(LPS/IFN-γ), and M(IL-4). See also Figures S2–S5 .

Article Snippet: Proteome profiler human protease/protease inhibitor arrays were purchased from R&D systems (Cat # ARY025).

Techniques: Cell Culture, Expressing, Comparison, Zymography, Enzyme-linked Immunosorbent Assay, Flow Cytometry

Journal: iScience

Article Title: Human monocyte-derived macrophages shift subcellular metalloprotease activity depending on their activation state

doi: 10.1016/j.isci.2024.111171

Figure Lengend Snippet:

Article Snippet: Proteome profiler human protease/protease inhibitor arrays were purchased from R&D systems (Cat # ARY025).

Techniques: Plasmid Preparation, Recombinant, Knock-Out, Blocking Assay, Staining, Antibody Labeling, Reverse Transcription, Lysis, Protease Inhibitor, Enzyme-linked Immunosorbent Assay, Fractionation, Cell Culture, Software, Flow Cytometry, Fluorescence, Spectrophotometry