anti eya2 rabbit polyclonal  (Proteintech)

Journal: International journal of molecular medicine

Article Title: Eya2 overexpression promotes the invasion of human astrocytoma through the regulation of ERK/MMP9 signaling.

doi: 10.3892/ijmm.2017.3132

Figure Lengend Snippet: Figure 1. Expression pattern of eyes absent (Eya) 2 in human astrocytoma. (A) Immunohistochemical staining of Eya2 protein in normal brain tissue was negative in glial cells and weak in neurons. The black arrows indicate neurons and the white arrows indicate glial cells. (B) Negative Eya2 staining in pilocytic astro- cytoma (grade I). (C) Negative Eya2 staining in grade II diffuse astrocytoma. (D) Weak nuclear Eya2 staining in grade III anaplastic astrocytoma. (E) Strong nuclear Eya2 staining in grade III anaplastic astrocytoma. (F) Strong nuclear Eya2 staining in glioblastoma (grade IV) (magnification, x200). Scale bar, 50 µm.

Article Snippet: The sections were deparaffinized and antigen retrieval was performed in citrate buffer for 2 min using an autoclave. endogenous peroxidase was then blocked using hydrogen peroxide. eya2 rabbit polyclonal antibody (1:300 dilution; 11314-1-aP; Proteintech inc., chicago, il, uSa) was used to incubate the tissue sections at 4̊C overnight.

Techniques: Expressing, Immunohistochemical staining, Staining

Journal: International journal of molecular medicine

Article Title: Eya2 overexpression promotes the invasion of human astrocytoma through the regulation of ERK/MMP9 signaling.

doi: 10.3892/ijmm.2017.3132

Figure Lengend Snippet: Figure 2. Eyes absent (Eya) 2 expression in astrocytoma cell lines. (A) Western blot analysis indicated that Eya2 and Six1 protein expression was high in the U251 cell line and low in the U87 and A172 cell lines. (B) Eya2 siRNA treatment decreased its mRNA and protein expression in U251 cells. *P<0.05 Eya2 siRNA vs Neg siRNA in U251 cells. Eya2 plasmid transfection upregulated its mRNA and protein levels. *P<0.05 Eya2 plasmid vs control plasmid in A172 cells.

Article Snippet: The sections were deparaffinized and antigen retrieval was performed in citrate buffer for 2 min using an autoclave. endogenous peroxidase was then blocked using hydrogen peroxide. eya2 rabbit polyclonal antibody (1:300 dilution; 11314-1-aP; Proteintech inc., chicago, il, uSa) was used to incubate the tissue sections at 4̊C overnight.

Techniques: Expressing, Western Blot, Plasmid Preparation, Transfection, Control

Journal: International journal of molecular medicine

Article Title: Eya2 overexpression promotes the invasion of human astrocytoma through the regulation of ERK/MMP9 signaling.

doi: 10.3892/ijmm.2017.3132

Figure Lengend Snippet: Figure 3. Eyes absent (Eya) 2 regulates the growth and invasion of astrocytoma cells. (A) Cell Counting kit-8 (CCK-8) assay demonstrated that Eya2 overex- pression accelerated cell growth rate, while Eya2 knockdown inhibited cell proliferation. (B) Colony formation assay demonstrated that Eya2 overexpres- sion upregulated the colony number, while Eya2 knockdown downregulated the colony number. *P<0.05 Eya2 siRNA vs Neg siRNA in U251 cells. *P<0.05 Eya2 plasmid vs control plasmid in A172 cells. (C) Matrigel invasion assay revealed that Eya2 knockdown decreased cell invasion in the U251 cell line. *P<0.05 Eya2 siRNA vs Neg siRNA in U251 cells. Transfection with Eya2 expession plasmid increased the A172 invading cell number. *P<0.05 Eya2 plasmid vs control plasmid in A172 cells.

Article Snippet: The sections were deparaffinized and antigen retrieval was performed in citrate buffer for 2 min using an autoclave. endogenous peroxidase was then blocked using hydrogen peroxide. eya2 rabbit polyclonal antibody (1:300 dilution; 11314-1-aP; Proteintech inc., chicago, il, uSa) was used to incubate the tissue sections at 4̊C overnight.

Techniques: Cell Counting, CCK-8 Assay, Knockdown, Colony Assay, Plasmid Preparation, Control, Invasion Assay, Transfection

Journal: International journal of molecular medicine

Article Title: Eya2 overexpression promotes the invasion of human astrocytoma through the regulation of ERK/MMP9 signaling.

doi: 10.3892/ijmm.2017.3132

Figure Lengend Snippet: Figure 4. Eyes absent (Eya) 2 regulates cell cycle and related protein expression. (A) Eya2 overexpression in A172 cells increased the S phase percentage and decreased G1 phase percentage. Eya2 knockdown by siRNA in U251 cells had the opposite effect. *P<0.05 Eya2 siRNA vs Neg siRNA in U251 cells. *P<0.05 Eya2 plasmid vs control plasmid in A172 cells. (B) Eya2 overexpression upregulated cyclin D1, cyclin E, MMP9 mRNA and protein expression in the A172 cell line. *P<0.05 Eya2 plasmid vs control plasmid in A172 cells. Eya2 knockdown downregulated the expression of these proteins in the U251 cells. *P<0.05 Eya2 siRNA vs Neg siRNA in U251 cells. Eya2 overexpression activated ERK phosphorylation, while Eya2 knockdown downregulated ERK phosphorylation.

Article Snippet: The sections were deparaffinized and antigen retrieval was performed in citrate buffer for 2 min using an autoclave. endogenous peroxidase was then blocked using hydrogen peroxide. eya2 rabbit polyclonal antibody (1:300 dilution; 11314-1-aP; Proteintech inc., chicago, il, uSa) was used to incubate the tissue sections at 4̊C overnight.

Techniques: Expressing, Over Expression, Knockdown, Plasmid Preparation, Control, Phospho-proteomics

Journal: International journal of molecular medicine

Article Title: Eya2 overexpression promotes the invasion of human astrocytoma through the regulation of ERK/MMP9 signaling.

doi: 10.3892/ijmm.2017.3132

Figure Lengend Snippet: Figure 5. Eyes absent (Eya) 2 interacts with Six1 and regulates ERK signaling. (A) ERK inhibitor, PD98059, blocked the promoting effect of Eya2 on A172 inva- sion. *P<0.05 Eya2 plasmid vs control plasmid in A172 cells. (B) Western blot analysis revealed that ERK inhibitor, PD98059, abolished the promoting effect of Eya2 on MMP9 mRNA and protein expression. *P<0.05 Eya2 plasmid vs control plasmid in A172 cells. (C) Eya2 co-immunoprecipitated with Six1 in U251 cells. (D) Transfection with Six1 siRNA significantly downregulated its protein in A172 cells. In Six1-depleted cells, transfection with Eya2 expression plasmid could not upregulate MMP9 expression at both the mRNA and protein level. *P<0.05 Eya2 plasmid vs control plasmid in A172 cells. Densitometry of western bands was measured using ImageJ software and relative intensity was calculated and indicated.

Article Snippet: The sections were deparaffinized and antigen retrieval was performed in citrate buffer for 2 min using an autoclave. endogenous peroxidase was then blocked using hydrogen peroxide. eya2 rabbit polyclonal antibody (1:300 dilution; 11314-1-aP; Proteintech inc., chicago, il, uSa) was used to incubate the tissue sections at 4̊C overnight.

Techniques: Plasmid Preparation, Control, Western Blot, Expressing, Immunoprecipitation, Transfection, Software