human tissue protein expression information  (Human Protein Atlas)

Journal: Biomolecules

Article Title: Expression of Periostin Alternative Splicing Variants in Normal Tissue and Breast Cancer

doi: 10.3390/biom14091093

Figure Lengend Snippet: Physiological distribution of Pn-ASV mRNA and total protein in adult mice. ( A ) The mPn-ASV mRNA copy number in several organs from C57BL6J mice aged at 8 weeks. Data are shown as mean ± SE, n = 3, * p < 0.05 vs. Pn 1–3. An amount of 1 μg total RNA was subjected to cDNA synthesis. The copy number of each Pn-ASV was calculated using specific primer sets and standard mouse Pn-ASV plasmid. ( B ) Total mPn protein expression in several organs from adult mice aged 8 weeks old. A total of 10 μg tissue proteins were loaded and detected with Pn exon 12 Ab, which detected all Pn-ASVs. A membrane with Coomassie brilliant blue (CBB) staining was shown as loading control. ( C ) Quantification data of total mPn protein expression in organs of adult mice.

Article Snippet: In addition, we referred to the human tissue protein expression information provided by the Human Protein Atlas ( www.proteinatlas.org (accessed on 10 November 2023)) [ ].

Techniques: cDNA Synthesis, Plasmid Preparation, Expressing, Membrane, Staining, Control

Journal: Biomolecules

Article Title: Expression of Periostin Alternative Splicing Variants in Normal Tissue and Breast Cancer

doi: 10.3390/biom14091093

Figure Lengend Snippet: Physiological distribution of mPn-ASV protein in adult mice. ( A ) Confirmation of total mPn protein expression in adult mouse organs. Total mPn protein expression was measured separately in each organ through immunoblotting with Pn exon 12 Ab. Supernatant from human Pn-1-Halotag over-expressed HEK293T cells (10 and 20 μg) and mouse 4T1 breast cancer cells (20 and 40 μg) were simultaneously blotted as a positive control. Proteins from Pn KO mice were used as a negative control. N = 3 for each organ. ( B ) IP and Western blotting results for mPn-ASVs. A total of 75 μg tissue proteins was immunoprecipitated with exon 14 (lane 2), 17 (lane 3), 21 (lane 4), and control IgG (lane 5). Immune complexes were collected on protein G-Agarose beads under agitation, according to the manufacturer’s instructions. Proteins were solubilized in Laemmli buffer, separated via SDS-PAGE, transferred to PVDF membranes, and detected with Pn exon 12 Ab. A total of 10 μg of un-precipitated sample was also loaded in order to determine the molecular level of physiologically expressed mPn-ASVs (lane 1). Lane 6 shows protein precipitated with beads only. The circle indicates the band of mPn-ASVs with exon 21. Coomassie brilliant blue (CBB) staining was performed, according to the manufacturer’s instructions. ( C ) Dosage-dependent increase in IP proteins of mPn-ASVs with exon 21. Amounts of 300, 150, and 75 μg of proteins were immunoprecipitated with Pn exon 21 Ab.

Article Snippet: In addition, we referred to the human tissue protein expression information provided by the Human Protein Atlas ( www.proteinatlas.org (accessed on 10 November 2023)) [ ].

Techniques: Expressing, Western Blot, Positive Control, Negative Control, Immunoprecipitation, Control, SDS Page, Staining

Journal: Biomolecules

Article Title: Expression of Periostin Alternative Splicing Variants in Normal Tissue and Breast Cancer

doi: 10.3390/biom14091093

Figure Lengend Snippet: Physiological distribution of hPn-ASV proteins in humans. ( A ) Total hPn expression in normal human organs. Tissue microarrays for normal human tissue were stained with Ab against n-terminal of hPn (Atlas Antibodies, Sigma-Aldrich, HPA012306). ( B ) IP and Western blotting for hPn-ASVs in normal stomach, colon, lung, and breast. A total of 75 μg of tissue proteins were immunoprecipitated with exon 14 (lane 2), 17 (lane 3), 21 (lane 4), and control IgG (lane 5). Immune complexes were collected on protein G-Agarose beads under agitation. Proteins were solubilized in Laemmli buffer, separated via SDS-PAGE, and transferred to PVDF membranes and detected with Pn exon 12 Ab. An amount of 10 μg of un-precipitated sample was also loaded to determine the molecular level of physiologically expressed Pn-ASVs (lane 1). Lane 6 shows beads only. The circle indicates the bands of hPn-ASVs with exon 21.

Article Snippet: In addition, we referred to the human tissue protein expression information provided by the Human Protein Atlas ( www.proteinatlas.org (accessed on 10 November 2023)) [ ].

Techniques: Expressing, Staining, Western Blot, Immunoprecipitation, Control, SDS Page

Journal: Biomolecules

Article Title: Expression of Periostin Alternative Splicing Variants in Normal Tissue and Breast Cancer

doi: 10.3390/biom14091093

Figure Lengend Snippet: hPn-ASVs expression in human breast cancer and adjacent normal tissue. ( A ) hPn-ASV mRNA expression in breast cancer (tumor) and normal adjacent tissue (NAT). Breast cancer specimens collected from patients undergoing total mastectomy for localized breast cancer were subjected to mRNA analysis. NAT was collected from the contralateral side of tumor across the nipple. Total hPn and hPn-ASVs with exon 17 or 21 were analyzed using specific primer sets and quantitative RT-PCR. Actual values for each patient ( A , B ) and average of hPn mRNA expression in tumor and NAT are also shown. ( C ) Data shown as mean ± SE, n = 11, * p < 0.05 vs. NAT. ( D ) hPn-ASV protein expression in tumor (T) and NAT (N) detected with Ab against exon 12 (all hPn-ASVs, left image) or 21 (right image). Immunoblot with exon 12 or 21 Ab was performed using 10 μg of proteins from 6 patients with breast cancer. ( E ) Quantitative analysis of change in expression in tumor with reference to NAT.

Article Snippet: In addition, we referred to the human tissue protein expression information provided by the Human Protein Atlas ( www.proteinatlas.org (accessed on 10 November 2023)) [ ].

Techniques: Expressing, Quantitative RT-PCR, Western Blot