anti-fgf8 antibody  (PeproTech)

Journal: bioRxiv

Article Title: Self-formation of concentric zones of telencephalic and ocular tissues and directional retinal ganglion cell axons

doi: 10.1101/2023.03.22.533827

Figure Lengend Snippet: A scheme of the procedure. (B) A diagram of CONCEPT organoids showing concentric zones of the anterior ectodermal progenitors. See also Figs.1, 2, 3, 5, S1, S10. (C) Morphology of cysts at day 2 showing the epithelial structure indicated by the apical reporter TJ::GFP at the lumen. (D) Morphology of CONCEPT organoids at day 26. (E-G) Expression of telencephalon (Tel) marker Foxg1, neuroretinal (NR) markers Vsx2 and Pax6 in mouse eyes at E10-10.5. Rostral optic stalk (OS) connected the telencephalic vesicle to the optic cup. (H-O) FOXG1+ telencephalic progenitors, VSX2+ and/or PAX6+ retinal progenitors formed concentric zones in CONCEPT organoids. N > 5 experiments. (P-W) In CONCEPT organoids, morphogens FGF8, BMP4, and BMP7 mRNA expression started at early stages and subsequently formed circular gradients. N > 5 experiments. Scale bars, 100 µm (C, E, M, O, P, S, T), 200 µm (I, K), 500 µm (Q, U), 1 mm (D, H, J, L, N, R, V, W).

Article Snippet: These primary antibodies were used: FOXG1 (Abcam, ab18259, 1:500), TUBB3 (Covance MMS-435P, 1:1000), FGF8 (1:500, R&D MAB323), RBPMS (1:200, PhosphoSolution 1830-RBPMS), ISL1 (1:500, DSHB 40.2D6), SNCG (1:200, Abcam ab55424), PAX2 (Invitrogen, 716000, 1:200), alpha A crystallin (Santa Cruz sc 22743, 1:500, shown as CRYAA in figure panels), beta crystallin (Santa Cruz sc-22745, 1:100, shown as CRY B in figure panels), CNTN2 (DSHB 4D7, 1:100), ALDH1A3 (Invitrogen, PA529188, 1:500), VAX1/2 (Santa Cruz sc-98613, 1:200), PAX6 (1:500, Covance PRB-278P), POU4F2 (Santa Cruz, SC-6026, 1:200), SIX3 (1:500, Rockland), and VSX2 (1:500, Millipore AB9016).

Techniques: Expressing, Marker

Journal: bioRxiv

Article Title: Self-formation of concentric zones of telencephalic and ocular tissues and directional retinal ganglion cell axons

doi: 10.1101/2023.03.22.533827

Figure Lengend Snippet: PAX2, FGF9, and FGF8 are differentially expressed in cluster 2, the major component of PAX2+ optic-disc cells. (B) PAX2 mRNA expression in CONCEPT organoids on day 25. Two PAX2+ concentric zones corresponding to the optic stalk (OS) and optic disc (OD) are labeled. (C) Dual-color immunocytochemistry indicates the co-localization of FGF8 and PAX2 in the optic-disc zone of CONCEPT organoids on day 25. (D-E) TUBB3+ axons grew towards and then along the cells that expressed high levels of FGF8 (D) and FGF9 mRNA (E) in CONCEPT organoids on day 25. Immunocytochemistry of TUBB3 was performed after in situ hybridization. (F-K) After the inhibition of FGF signaling with FGFR inhibitor PD 161570 during days 17-24, the number of RGC somas drastically reduced, and directional axon growth of RGCs was nearly absent, whereas FGF8 protein expression largely remained. CNTN2 immunocytochemistry before (F, H) and after FGF8 immunocytochemistry (G, I, J, K) are shown. N = 3/3 experiments. Scale bar, 250 µm (B), 100 µm (C-E), 1 mm (F), 200 µm (K).

Article Snippet: These primary antibodies were used: FOXG1 (Abcam, ab18259, 1:500), TUBB3 (Covance MMS-435P, 1:1000), FGF8 (1:500, R&D MAB323), RBPMS (1:200, PhosphoSolution 1830-RBPMS), ISL1 (1:500, DSHB 40.2D6), SNCG (1:200, Abcam ab55424), PAX2 (Invitrogen, 716000, 1:200), alpha A crystallin (Santa Cruz sc 22743, 1:500, shown as CRYAA in figure panels), beta crystallin (Santa Cruz sc-22745, 1:100, shown as CRY B in figure panels), CNTN2 (DSHB 4D7, 1:100), ALDH1A3 (Invitrogen, PA529188, 1:500), VAX1/2 (Santa Cruz sc-98613, 1:200), PAX6 (1:500, Covance PRB-278P), POU4F2 (Santa Cruz, SC-6026, 1:200), SIX3 (1:500, Rockland), and VSX2 (1:500, Millipore AB9016).

Techniques: Expressing, Labeling, Immunocytochemistry, In Situ Hybridization, Inhibition

Journal: Oncology Letters

Article Title: Systematic analysis of expression profiles and prognostic significance of the FGF gene family in pancreatic adenocarcinoma

doi: 10.3892/ol.2022.13555

Figure Lengend Snippet: Expression of FGFs in PAAD (Gene Expression Profiling Interactive Analysis). Expression of (A) FGF2, (B) FGF8, (C) FGF9, (D) FGF13, (E) FGF17 and (F) FGF22 in PAAD. *P<0.05. FGF, fibroblast growth factor; PAAD, pancreatic adenocarcinoma; N, normal tissues; T, tumor tissues.

Article Snippet: A total of 50 μl of 5–10% normal goat serum was added per chip for blocking (1:19 fold dilution) at room temperature for 30 min. Immunohistochemical staining of the paraffin-embedded tissues was performed using FGF2 (1:200; sc-74412; Santa Cruz Biotechnology, Inc.) and FGF8 (1:200; 20711-1-AP; ProteinTech Group, Inc.) primary antibodies, anti-mouse secondary antibodies (1:200; GB23301; Wuhan Servicebio Technology Co., Ltd.), anti-rabbit secondary antibodies (1:200; GB23303; Wuhan Servicebio Technology Co., Ltd.), and an ABC Elite immunoperoxidase kit (Wuhan Servicebio Technology Co., Ltd.) according to the manufacturer's instructions.

Techniques: Expressing, Gene Expression

Journal: Oncology Letters

Article Title: Systematic analysis of expression profiles and prognostic significance of the FGF gene family in pancreatic adenocarcinoma

doi: 10.3892/ol.2022.13555

Figure Lengend Snippet: Prognostic value of FGFs in PAAD (Gene Expression Profiling Interactive Analysis). Prognostic value of the mRNA expression of (A) FGF2, (B) FGF8, (C) FGF9, (D) FGF13, (E) FGF17 and (F) FGF22 in PAAD. FGF, fibroblast growth factor; PAAD, pancreatic adenocarcinoma; TPM, transcripts per million.

Article Snippet: A total of 50 μl of 5–10% normal goat serum was added per chip for blocking (1:19 fold dilution) at room temperature for 30 min. Immunohistochemical staining of the paraffin-embedded tissues was performed using FGF2 (1:200; sc-74412; Santa Cruz Biotechnology, Inc.) and FGF8 (1:200; 20711-1-AP; ProteinTech Group, Inc.) primary antibodies, anti-mouse secondary antibodies (1:200; GB23301; Wuhan Servicebio Technology Co., Ltd.), anti-rabbit secondary antibodies (1:200; GB23303; Wuhan Servicebio Technology Co., Ltd.), and an ABC Elite immunoperoxidase kit (Wuhan Servicebio Technology Co., Ltd.) according to the manufacturer's instructions.

Techniques: Gene Expression, Expressing

Journal: Oncology Letters

Article Title: Systematic analysis of expression profiles and prognostic significance of the FGF gene family in pancreatic adenocarcinoma

doi: 10.3892/ol.2022.13555

Figure Lengend Snippet: Correlations between differentially expressed FGFs and immune cell infiltration (Tumor Immune Estimation Resource). Correlations between the abundance of immune cells and the expression of (A) FGF2, (B) FGF8, (C) FGF9, (D) FGF13, (E) FGF17 and (F) FGF22. FGF, fibroblast growth factor; PAAD, pancreatic adenocarcinoma; TPM, transcripts per million.

Article Snippet: A total of 50 μl of 5–10% normal goat serum was added per chip for blocking (1:19 fold dilution) at room temperature for 30 min. Immunohistochemical staining of the paraffin-embedded tissues was performed using FGF2 (1:200; sc-74412; Santa Cruz Biotechnology, Inc.) and FGF8 (1:200; 20711-1-AP; ProteinTech Group, Inc.) primary antibodies, anti-mouse secondary antibodies (1:200; GB23301; Wuhan Servicebio Technology Co., Ltd.), anti-rabbit secondary antibodies (1:200; GB23303; Wuhan Servicebio Technology Co., Ltd.), and an ABC Elite immunoperoxidase kit (Wuhan Servicebio Technology Co., Ltd.) according to the manufacturer's instructions.

Techniques: Expressing

Journal: Oncology Letters

Article Title: Systematic analysis of expression profiles and prognostic significance of the FGF gene family in pancreatic adenocarcinoma

doi: 10.3892/ol.2022.13555

Figure Lengend Snippet: Expression and influence of FGF2 and FGF8 in pancreatic adenocarcinoma. (A) Expression of FGF2 and FGF8 in pancreatic adenocarcinoma. Scale bar, 20 µm. Quantification analysis of IHC for (B) FGF2 (P=0.0313; Wilcoxon's signed rank test) and (C) FGF8 (P=0.0391; Wilcoxon's signed rank test) (*P<0.05). (D) Colony formation assay using MIA Paca-2 cells treated with DMSO or 10 µM alofanib for 7 days. (E) Colony numbers are presented as the mean ± SD (n=3). *P<0.05. Unpaired Student's t-test. FGF, fibroblast growth factor; IHC, immunohistochemistry; N, normal tissue; T, tumor tissue.

Article Snippet: A total of 50 μl of 5–10% normal goat serum was added per chip for blocking (1:19 fold dilution) at room temperature for 30 min. Immunohistochemical staining of the paraffin-embedded tissues was performed using FGF2 (1:200; sc-74412; Santa Cruz Biotechnology, Inc.) and FGF8 (1:200; 20711-1-AP; ProteinTech Group, Inc.) primary antibodies, anti-mouse secondary antibodies (1:200; GB23301; Wuhan Servicebio Technology Co., Ltd.), anti-rabbit secondary antibodies (1:200; GB23303; Wuhan Servicebio Technology Co., Ltd.), and an ABC Elite immunoperoxidase kit (Wuhan Servicebio Technology Co., Ltd.) according to the manufacturer's instructions.

Techniques: Expressing, Colony Assay, Immunohistochemistry

Journal: OncoImmunology

Article Title: Immunoreactivity against fibroblast growth factor 8 in alveolar rhabdomyosarcoma patients and its involvement in tumor aggressiveness

doi: 10.1080/2162402x.2022.2096349

Figure Lengend Snippet: Figure 1. FGF8 autoantibodies detection in very high-risk ARMS patients. (a) Left, experimental workflow followed for the identification of autoantibodies in ARMS patients using plasma samples and ProtoArrayTM technology.31 The immune response profile was obtained from 10 metastatic ARMS patients and 15 healthy subjects (HS), probing protein microarray chips with plasma. Reactivity of 9374 spotted antigens was evaluated after signal detection, filtration and normalization using robust linear model (RLM). Antigens median values were calculated for each group, compared and ranked according to significant p-value scale. Right, volcano plot of protein microarray data showing differentially immunoreactive antigens between patients and healthy subjects, plotted along dimension of fold change (abscissae) and statistical difference (ordinates). Antigens with significant p-values (≥0,05) are indicated by colors and names, while antigens with no significant difference in immunoreactivity between patients and controls are plotted uncolored on the bottom of the graph (gray dots). Antigens more reactive in patients or controls are distinguished by red or green dots, respectively. (b) Venn diagram showing the overlap between differential immunoreactive antigens (n = 55) and PAX3-FOXO1 target genes (n = 1010).32 (c) Box plot of FGF8 signal intensity revealed in patients and controls by protein microarray (RFU = relative fluorescence unit) and (d) validation by indirect ELISA assay. (e) Correlation of FGF8 autoantibody signal intensity and FGF8 IgG concentration obtained in the same samples cohort by protein microarrays and ELISA assay, respectively. p < 0,05 (*); p < 0,01 (**).

Article Snippet: Primary antibody against FGF8 (Proteintech) and in 1%BSA/PBS1X were probed at 37°C for 60 minutes, followed by secondary antibody Alexa Fluor® 546 conjugate (Thermo Fisher scientific) in PBS1X at 37°C for 60 minutes.

Techniques: Clinical Proteomics, Microarray, Filtration, Fluorescence, Biomarker Discovery, Indirect ELISA, Concentration Assay, Enzyme-linked Immunosorbent Assay

Journal: OncoImmunology

Article Title: Immunoreactivity against fibroblast growth factor 8 in alveolar rhabdomyosarcoma patients and its involvement in tumor aggressiveness

doi: 10.1080/2162402x.2022.2096349

Figure Lengend Snippet: Figure 2. Correlation between humoral immune response against FGF8 and patients’ outcome. Kaplan-Meier survival analysis representing (a) event-free survival (EFS) and (b) overall survival (OS) of ARMS patients distinguished accord ing to FGF8 autoantibodies median value.

Article Snippet: Primary antibody against FGF8 (Proteintech) and in 1%BSA/PBS1X were probed at 37°C for 60 minutes, followed by secondary antibody Alexa Fluor® 546 conjugate (Thermo Fisher scientific) in PBS1X at 37°C for 60 minutes.

Techniques:

Journal: OncoImmunology

Article Title: Immunoreactivity against fibroblast growth factor 8 in alveolar rhabdomyosarcoma patients and its involvement in tumor aggressiveness

doi: 10.1080/2162402x.2022.2096349

Figure Lengend Snippet: Figure 3. Expression of FGF8 in RMS cell lines. (a) Relative quantification of FGF8 mRNA by qRT-PCR in normal control cells (CTR, n = 4), alveolar RMS (ARMS, n = 5), embryonal RMS (ERMS, n = 5), Ewing sarcoma (EWS, n = 7), Non-Hodgkin lymphoma cell lines (NHL, n = 5), leukemia cell lines (Leukemia, n = 3) and cell lines of various origins (Others, n = 10). Statistical significance was calculated by Mann-Whitney U test, comparing each group of cell lines with ARMS group. Glyceraldehyde- 3-phosphate dehydrogenase (GAPDH) housekeeping gene was used for normalization, while normal skeletal muscle tissue extracts were used as external calibrator. (b) FGF8 protein expression and localization by immunocytochemistry and (c) immunofluorescence analysis in RH30 (PF+ ARMS) and RD (PF− ERMS) cell lines (magnifications of selected areas are shown apart). (d) Western blotting and (e) direct ELISA assay performed using serum-starved RH30 and RD total lysate and growth medium, respectively, to assess FGF8 protein at intracellular and at secreted level. A_SKM, adult skeletal muscle; p < 0,05 (*); p < 0,01 (**).

Article Snippet: Primary antibody against FGF8 (Proteintech) and in 1%BSA/PBS1X were probed at 37°C for 60 minutes, followed by secondary antibody Alexa Fluor® 546 conjugate (Thermo Fisher scientific) in PBS1X at 37°C for 60 minutes.

Techniques: Expressing, Quantitative Proteomics, Quantitative RT-PCR, Control, MANN-WHITNEY, Immunocytochemistry, Immunofluorescence, Western Blot, Direct ELISA

Journal: OncoImmunology

Article Title: Immunoreactivity against fibroblast growth factor 8 in alveolar rhabdomyosarcoma patients and its involvement in tumor aggressiveness

doi: 10.1080/2162402x.2022.2096349

Figure Lengend Snippet: Figure 4. FGF8 signaling in RMS cells. (a) Relative quantification of FGFR1-4 receptors mRNA in ARMS and ERMS cell lines. FGF8 mRNA levels are also displayed in graph (red open dots). Glyceraldehyde- 3-phosphate dehydrogenase (GAPDH) was used for signal intensity normalization, while skeletal muscle extracts were used as external calibrator. (b) FGFR2 and FGFR4 expression and phosphorylation in PF+ (RH30, RH4) and PF− (RD) cell lines, performed through immunoprecipitation of total FGFR2 and FGFR4 receptor proteins. GAPDH protein was used as gel-loading control. (c) Western blotting showing time-dependent phosphorylation of FRS2 and ERK1/2 proteins induced in RH4 and RD cells upon exposure to 50 ng/mL human recombinant FGF8 protein for the indicated time periods. Between blots a graph displaying phosphorylated FRS2 and ERK1/2 band densities, quantified using ImageJ software, was included. γ-Tubulin was used as gel loading control. (d) RH4 and RD cell lines wound healing assay performed in presence and absence of 100 ng/ml human FGF8. Images were taken up to 48 hours after the treatment. (e) Immunoblot analysis of phosphorylated ERK1/2 kinase in RMS cells exposed to increasing concentration of human FGF8 (25, 100 ng/mL), pretreated or not for 2 hours with 5 μM of NVP-BGJ398. γ-Tubulin was used as gel loading control. (f) MTT assay showing PF+ ARMS (RH30, RH4) and PF− ERMS (RH36, RD) cell viability in the presence of 5 μM of NVP-BGJ398 up to 72 hours.

Article Snippet: Primary antibody against FGF8 (Proteintech) and in 1%BSA/PBS1X were probed at 37°C for 60 minutes, followed by secondary antibody Alexa Fluor® 546 conjugate (Thermo Fisher scientific) in PBS1X at 37°C for 60 minutes.

Techniques: Quantitative Proteomics, Expressing, Phospho-proteomics, Immunoprecipitation, Control, Western Blot, Recombinant, Software, Wound Healing Assay, Concentration Assay, MTT Assay

Journal: OncoImmunology

Article Title: Immunoreactivity against fibroblast growth factor 8 in alveolar rhabdomyosarcoma patients and its involvement in tumor aggressiveness

doi: 10.1080/2162402x.2022.2096349

Figure Lengend Snippet: Figure 5. FGF8-induced gene expression in RMS cells. (a) Time-dependent expression of DUSP6, SPRY4, GDF15 and ETV5 FGF-target genes, upon treatment of RH4 and RD cells with 100 ng/ml of human recombinant FGF8. Glyceraldehyde- 3-phosphate dehydrogenase (GAPDH) housekeeping gene was used for normalization. (b) STRING analysis. (c) Time-dependent expression of PLAU and MMP-9 genes after treatment of RH4 and RD cell lines with 100 ng/ml of human recombinant FGF8.

Article Snippet: Primary antibody against FGF8 (Proteintech) and in 1%BSA/PBS1X were probed at 37°C for 60 minutes, followed by secondary antibody Alexa Fluor® 546 conjugate (Thermo Fisher scientific) in PBS1X at 37°C for 60 minutes.

Techniques: Gene Expression, Expressing, Recombinant

Journal: OncoImmunology

Article Title: Immunoreactivity against fibroblast growth factor 8 in alveolar rhabdomyosarcoma patients and its involvement in tumor aggressiveness

doi: 10.1080/2162402x.2022.2096349

Figure Lengend Snippet: Figure 6. FGF8 expression in RMS tumor biopsies. (a) Relative quantification of FGF8 mRNA in RMS primary tumors (n = 50) and normal controls (n = 4), and (b) in RMS primary tumors divided according to fusion status. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene was used for normalization. (c) Hematoxylin/Eosin (HE) and FGF8 staining of representative ARMS and ERMS cases. (d) Scatter plot showing the correlation between FGF8 mRNA levels and autoantibodies titer in 33 PF+ ARMS primary tumors and plasma samples, respectively. Vertical and horizontal dashed lines represent FGF8 mRNA and autoantibody median values, respectively, used to divide the plot in four regions (I–IV). Dots represent patients, labeled with different colors based on event occurrence (gray) or not (red) after frontline chemotherapy. p < 0,001 (***); p < 0,0001 (****).

Article Snippet: Primary antibody against FGF8 (Proteintech) and in 1%BSA/PBS1X were probed at 37°C for 60 minutes, followed by secondary antibody Alexa Fluor® 546 conjugate (Thermo Fisher scientific) in PBS1X at 37°C for 60 minutes.

Techniques: Expressing, Quantitative Proteomics, Staining, Clinical Proteomics, Labeling

Journal: OncoImmunology

Article Title: Immunoreactivity against fibroblast growth factor 8 in alveolar rhabdomyosarcoma patients and its involvement in tumor aggressiveness

doi: 10.1080/2162402x.2022.2096349

Figure Lengend Snippet: Figure 7. FGF8 expression in RMS recurrent tumors. (a) FGF8 and PAX3-FOXO1 or PAX7-FOXO1, mRNA at diagnosis and at relapse in 7 cases of ARMS, and (b) 5 ERMS cases. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) gene was used for normalization. n.s., not significant; p < 0,05 (*).

Article Snippet: Primary antibody against FGF8 (Proteintech) and in 1%BSA/PBS1X were probed at 37°C for 60 minutes, followed by secondary antibody Alexa Fluor® 546 conjugate (Thermo Fisher scientific) in PBS1X at 37°C for 60 minutes.

Techniques: Expressing, Biomarker Discovery