recombinant fgf4  (R&D Systems)

Journal: Genes & Development

Article Title: Transcriptional derepression of negative regulators of MAP kinase supports maintenance of diapause ES cells in the pluripotent state

doi: 10.1101/gad.353143.125

Figure Lengend Snippet: The differentiation block in I-BET-resistant (I-BETR) diapause-like ES cells. ( A ) Alkaline phosphatase (AP) levels in control and I-BETR ES cells. Scale bar, 100 µm. ( B ) Expression levels of the pluripotency ( left panel) or differentiation-inducing ( right panel) genes in control, 2i-treated, or I-BETR ES cells. RNA expression levels were quantified by qPCR. Values represent normalized mean ± SD. n = 3. ( C ) Colony morphology and/or alkaline phosphatase (AP) levels in control and I-BETR ES cells following vehicle or FGF4 triggering. Scale bar, 100 µm. The bar graph represents quantification of relative percentage of pluripotent colonies in different groups. Values represent ± SD. n = 9. (n.s.) No significance, (****) P < 0.0001, one-way ANOVA with Dunnett's multiple comparisons test. ( D ) Expression levels of selected pluripotency ( left ) or differentiation-inducing ( right ) genes in control and I-BETR ES cells treated or not treated with FGF4. Error bars indicate SD. n = 3. (n.s.) No significance, (****) P < 0.0001, one-way ANOVA with Dunnett's multiple comparisons test. ( E ) Withdrawal of I-BET (I-BETW) restores the control ES cell-like pluripotency gene expression pattern in I-BETR ES cells. Values represent gene expression levels normalized to the mean of control samples based on TPM values observed by bulk mRNA RNA-seq analysis . ( F ) Generation of chimeras by I-BETR ES cells. The I-BETR ES cells were incubated in I-BET-free medium for 12–14 h and injected into the C57BL/6J blastocysts. White coat color indicates the chimerism.

Article Snippet: For FGF4-driven ES cell differentiation, 10 ng/μL recombinant FGF4 (R&D Systems 235-F4) was added on day 0 together with 1 μg/μL heparin (Sigma-Aldrich H3149).

Techniques: Blocking Assay, Control, Expressing, RNA Expression, Gene Expression, RNA Sequencing, Incubation, Injection

Journal: Antibodies

Article Title: Two Highly Specific Mouse Monoclonal Antibodies to the Putative C-Telopeptide of Human Collagen XIα1, a Cancer Biomarker

doi: 10.3390/antib15020021

Figure Lengend Snippet: The amino acid sequence of the COL11A1 Fusion Protein (residues 1545 to 1806 of the P12107-1 A isoform), recombinantly expressed in Escherichia coli , after the removal of an N-terminal GST tag (provided by Proteintech). The first 19 N-terminal (1545) PLPILSSKKTRRHTEGMQA (1563) amino acid residues are part of the putative C-telopeptide. The next 243 amino acid residues (1564 to 1806) constitute the C-propeptide.

Article Snippet: The recombinant COL11A1 Fusion Protein from Proteintech was also assayed in an ELISA and Western blot with finally purified preparations of the anti-colXIα1 clone 9 and PLY-7 mAbs. shows their ELISA immunoreactivity characteristics.

Techniques: Sequencing

Journal: Antibodies

Article Title: Two Highly Specific Mouse Monoclonal Antibodies to the Putative C-Telopeptide of Human Collagen XIα1, a Cancer Biomarker

doi: 10.3390/antib15020021

Figure Lengend Snippet: SDS-PAGE gel staining and Western blot of recombinant antigens with finally purified preparations of anti-colXIα1 clone 9 and PLY-7 mAbs. Lane 1: PageRuler™ Plus Prestained Protein Ladder. Lane 2: COL11A1 Fusion Protein (Proteintech). Lane 3: Collagen XIα1 (GenScript). Lane 4: C-propeptide (GenScript). Western blot color development was monitored following the manufacturer’s instructions. Full-length blots/gels are presented in .

Article Snippet: The recombinant COL11A1 Fusion Protein from Proteintech was also assayed in an ELISA and Western blot with finally purified preparations of the anti-colXIα1 clone 9 and PLY-7 mAbs. shows their ELISA immunoreactivity characteristics.

Techniques: SDS Page, Staining, Western Blot, Recombinant, Purification

Journal: Antibodies

Article Title: Two Highly Specific Mouse Monoclonal Antibodies to the Putative C-Telopeptide of Human Collagen XIα1, a Cancer Biomarker

doi: 10.3390/antib15020021

Figure Lengend Snippet: The PEP-FOLD4-derived structural predictions of peptides related to the putative C-telopeptide. ( A ): The 50 N-terminal amino acid sequence of the COL11A1 Fusion Protein from Proteintech, with the first 19 N-terminal PLPILSSKKTRRHTEGMQA amino acid residues of the putative C-telopeptide. ( B ): A free RRHTEGMQA peptide. ( C ): The 50 C-terminal amino acid sequence of GenScript’s recombinant collagen XIα1 form, whose last 21 C-terminal IQPLPILSSKKTRRHTEGMQA amino acid residues correspond to the putative C-telopeptide. The peptide’s N-terminus is on the left in ( A , B ) and on the right in ( C ).

Article Snippet: The recombinant COL11A1 Fusion Protein from Proteintech was also assayed in an ELISA and Western blot with finally purified preparations of the anti-colXIα1 clone 9 and PLY-7 mAbs. shows their ELISA immunoreactivity characteristics.

Techniques: Derivative Assay, Sequencing, Recombinant

Journal: Advanced Science

Article Title: Mutual Exclusion Analysis Shows that DUSP9 Negatively Regulates PD‐L1 Expression and Acts as a Target to Enhance Anti‐PD‐1 Efficacy

doi: 10.1002/advs.202514347

Figure Lengend Snippet: Identification of the mutually exclusive molecules of PD‐L1 expression. A,B) A model showing unilateral exclusion due to the low plasticity of PD‐L1 (A) and any of the other genes (B), such as gene i . Each point represents a sample. C) A model of bilateral exclusion is used in this study. D) Linear negative correlation comprises co‐expression with relatively high expression levels in the indicated samples. E) Cosine similarity is sensitive to outlier values. F) The workflow chart shows the PD‐L1 mutually exclusive genes screening by virtual sorting. G) The volcano plot shows the delta value and adjusted p value of differential expression based on virtual sorting of PD‐L1 high and PD‐L1 low/− samples. H) The Venn diagram shows the number of genes that intersect across three screening conditions: cosine ≤ 0.05, delta ≤ ‐15, and adjusted p value ≤ 0.01. I) The expression of CORO1A, DUSP9, FGFR3, FGFR4, and KCNH2 was examined using A549 RNA‐Seq data. Data are shown as mean ± SEM; n = 2; adjusted p values were calculated by DESeq2. J,K) The expression of PD‐L1 with the indicated gene knockdown was detected by qPCR in A549 (J) and Huh7 (K) cells. Data are shown as mean ± SEM; n = 3; p values were determined by Student's t‐test; ns, not significant; * , p < 0.05; ** , p < 0.01; *** , p < 0.001; ****, p < 0.0001. L) Survival analysis of patients who received anti‐PD‐1 therapy, grouped by CD274 and the indicated gene expression, as determined by Kaplan‐Meier Plotter (http://kmplot.com/analysis/).

Article Snippet: Human recombinant DUSP9 (Cat#AG5881) was purchased from Proteintech.

Techniques: Expressing, Quantitative Proteomics, RNA Sequencing, Knockdown, Gene Expression

Journal: Advanced Science

Article Title: Mutual Exclusion Analysis Shows that DUSP9 Negatively Regulates PD‐L1 Expression and Acts as a Target to Enhance Anti‐PD‐1 Efficacy

doi: 10.1002/advs.202514347

Figure Lengend Snippet: DUSP9 negatively regulates PD‐L1 expression in tumor cells. A–C) The expression of PD‐L1 in DUSP9‐overexpression (DUSP9‐OE) tumor cell lines, including A549 (A), FaDu (B), and HeLa (C), was examined by Western blot. D‐F) The expression of PD‐L1 in DUSP9‐knockdown (DUSP9‐KD) Huh7 cells was detected by Western blot (D), flow cytometry (E), and qPCR (F). G‐I) Flow cytometry results show the expression of PD‐L1 in DUSP9‐KD Hepa1‐6 cells (G), MC38 cells (H), and LLC cells (I). Data are shown as mean ± SEM; n = 3; p values were determined by two‐way ANOVA with Sidak's (A–C) or Dunnett's (D, F) post‐hoc test; ns, not significant; ** , p < 0.01; *** , p < 0.001; **** , p < 0.0001.

Article Snippet: Human recombinant DUSP9 (Cat#AG5881) was purchased from Proteintech.

Techniques: Expressing, Over Expression, Western Blot, Knockdown, Flow Cytometry

Journal: Advanced Science

Article Title: Mutual Exclusion Analysis Shows that DUSP9 Negatively Regulates PD‐L1 Expression and Acts as a Target to Enhance Anti‐PD‐1 Efficacy

doi: 10.1002/advs.202514347

Figure Lengend Snippet: DUSP9 reduces PD‐L1 expression by catalyzing the dephosphorylation of STAT3. A) The abundances of DUSP9 and STAT3 were compared between the IgG and anti‐DUSP9 groups using IP‐MS. B) Western blot results show the expression of PD‐L1 and the phosphorylation of STAT3 in DUSP9‐KD Huh7 cells. C) Immunofluorescence results show the phosphorylation of STAT3 in DUSP9‐KD Huh7 cells. D) The expression of PD‐L1 and the phosphorylation of STAT3 in Huh7 cells after DUSP9 and STAT3 knockdown were examined by Western blot. E) The expression of PD‐L1 and the phosphorylation of STAT3 in DUSP9‐KD Huh7 cells treated with the p‐STAT3‐S727‐specific inhibitor, S3I‐201, were examined by Western blot. F,G) Co‐IP results show the endogenous interaction between DUSP9 and STAT3 when using DUSP9 antibody (F) and STAT3 antibody (G). H) In vitro dephosphorylation assay shows the direct dephosphorylating function of DUSP9 on p‐STAT3 at both Y705 and S727. Positive control: calf intestinal alkaline phosphatase (CIAP). Data are shown as mean ± SEM; n = 3; p values were determined by Student's t‐test (A), two‐way ANOVA with Dunnett's post‐hoc test (B), two‐way ANOVA with Sidak's (for PD‐L1) or Tukey's (for p‐STAT3) post‐hoc test (D, E), and one‐way ANOVA with Dunnett's post‐hoc test (H); ns, not significant; * , p < 0.05; ** , p < 0.01; *** , p < 0.001; **** , p < 0.0001.

Article Snippet: Human recombinant DUSP9 (Cat#AG5881) was purchased from Proteintech.

Techniques: Expressing, De-Phosphorylation Assay, Protein-Protein interactions, Western Blot, Phospho-proteomics, Immunofluorescence, Knockdown, Co-Immunoprecipitation Assay, In Vitro, Positive Control

Journal: Advanced Science

Article Title: Mutual Exclusion Analysis Shows that DUSP9 Negatively Regulates PD‐L1 Expression and Acts as a Target to Enhance Anti‐PD‐1 Efficacy

doi: 10.1002/advs.202514347

Figure Lengend Snippet: DUSP9 knockdown inhibits tumor growth in vitro and in vivo. A) Colony formation assays were conducted on DUSP9‐KD Hepa1‐6 cells, MC38 cells, and LLC cells. Data are shown as mean ± SEM; n = 2; p values were determined by two‐way ANOVA with Dunnett's post‐hoc test; * , p < 0.05; ** , p < 0.01. B–J) An overview of the subcutaneous transplantation tumors of DUSP9‐KD Hepa1‐6 cells (B), MC38 cells (E), and LLC cells (H) in BALB/c nude mice. Tumor growth (C, F, I) and tumor weight (D, G, J) are also shown. Data are shown as mean ± SEM; n = 4 (B–G) or 6 (H–J); p values were determined by Student's t‐test; ** , p < 0.01; *** , p < 0.001.

Article Snippet: Human recombinant DUSP9 (Cat#AG5881) was purchased from Proteintech.

Techniques: Knockdown, In Vitro, In Vivo, Transplantation Assay

Journal: Advanced Science

Article Title: Mutual Exclusion Analysis Shows that DUSP9 Negatively Regulates PD‐L1 Expression and Acts as a Target to Enhance Anti‐PD‐1 Efficacy

doi: 10.1002/advs.202514347

Figure Lengend Snippet: DUSP9 knockdown inhibits cytotoxic CD8 + T cells by inducing PD‐L1 expression. A–E) The syngeneic tumor model of DUSP9‐KD MC38 cells in C57BL/6 mice includes the following: (A) tumor overview; (B) tumor growth; (C) tumor weight; (D) statistics on the percentage of tumor‐infiltrating T cell subsets; (E) GZMB + CD8 + T cell and TNF‐α + CD8 + T cell subsets detected by flow cytometry. F–H) Co‐culture of primary T cells and DUSP9‐KD Hepa1‐6 cells with anti‐PD‐1 treatment: (F) flowchart; (G) GZMB + CD8 + T cell subset detected by flow cytometry; (H) statistics on the percentage of the GZMB + CD8 + T cell subset. Data are shown as mean ± SEM; n = 6 (A–D) or 4 (FH); p values were determined by Student's t ‐test (B–D) or two‐way ANOVA with Dunnett's post‐hoc test (H); ns, not significant; * , p < 0.05; ** , p < 0.01; *** , p < 0.001.

Article Snippet: Human recombinant DUSP9 (Cat#AG5881) was purchased from Proteintech.

Techniques: Knockdown, Expressing, Flow Cytometry, Co-Culture Assay

Journal: Advanced Science

Article Title: Mutual Exclusion Analysis Shows that DUSP9 Negatively Regulates PD‐L1 Expression and Acts as a Target to Enhance Anti‐PD‐1 Efficacy

doi: 10.1002/advs.202514347

Figure Lengend Snippet: DUSP9 knockdown promotes antitumor efficacy in combination with anti‐PD‐1 therapy. The syngeneic tumor model of C57BL/6 mice was inoculated with various cells and treated as follows: A) DUSP9‐KD MC38 cells and treated with anti‐PD‐1; B) DUSP9‐KD LLC cells and treated with anti‐PD‐1; C) wild‐type MC38 cells and treated with si Dusp9 and anti‐PD‐1; D) wild‐type LLC cells and treated with si Dusp9 and anti‐PD‐1. Data are shown as mean ± SEM; n = 6; p values were determined by one‐way ANOVA with Sidak's post‐hoc test; ns, not significant; * , p < 0.05; ** , p < 0.01; *** , p < 0.001; **** , p < 0.0001.

Article Snippet: Human recombinant DUSP9 (Cat#AG5881) was purchased from Proteintech.

Techniques: Knockdown

Journal: Advanced Science

Article Title: Mutual Exclusion Analysis Shows that DUSP9 Negatively Regulates PD‐L1 Expression and Acts as a Target to Enhance Anti‐PD‐1 Efficacy

doi: 10.1002/advs.202514347

Figure Lengend Snippet: DUSP9 may serve as a clinical predictor of ICB therapy efficacy. A–C) The mRNA expression of DUSP9 and PD‐L1 between responders and non‐responders in various tumors was investigated: (A) melanoma treated with anti‐PD‐L1 therapy (reanalysis of the GSE172320 dataset); (B) mesothelioma treated with anti‐CTLA4 and anti‐PD‐L1 therapy (reanalysis of the GSE117358 dataset); (C) HCC treated with anti‐PD‐L1 therapy (reanalysis of the GSE279750 dataset). The scatter plots show the correlation between the mRNA expression levels of DUSP9 and PD‐L1 according to each dataset. D) Immunohistochemistry results show the expression of DUSP9 in responders and non‐responders to anti‐PD‐1/PD‐L1 therapy among clinical HNSC patients. Data are shown as mean ± SEM; p values were determined by Student's t‐test (A, B, D) or Mann‐Whitney test (C); ns, not significant; ** , p < 0.01; **** , p < 0.0001.

Article Snippet: Human recombinant DUSP9 (Cat#AG5881) was purchased from Proteintech.

Techniques: Expressing, Immunohistochemistry, MANN-WHITNEY