rabbit monoclonal anti-vitronectin  (Novus Biologicals)

Journal: Therapeutic Advances in Urology

Article Title: The role of chromodomain helicase DNA binding protein 1 (CHD1) in promoting an invasive prostate cancer phenotype

doi: 10.1177/17562872211022462

Figure Lengend Snippet: Expression of extracellular matrix proteins and adhesion molecules in CHD1 KO cells. (a) Graph demonstrating the expression of genes that were altered at least two-fold in the NT2 and CHD1 KO lines, Cr16 and Cr21, compared to RWPE-1. ITGA2 was down-regulated, while ITGA4 is up-regulated in CHD1 KO cells. The integrin ligands, collagen (COL16A1, COL4A2, COL5A1 and COL6A2), FN1, and the laminin component, LAMB3, are downregulated in CHD1 KO cells. The most down-regulated ECM components in the CHD1 KO cells were MMP2, SPARC and VTN. The ECM proteins ITGA4, MMP12, and SELL were up regulated. (b) Plot depicting levels of secreted SPARC protein as detected by ELISA. High levels of SPARC are secreted by RWPE-1 compared to the CHD1 KO lines ( ** p < 0.01). NT2 cell lines secreted higher levels of SPARC than RWPE-1 cell lines ( * p < 0.05). Error bars represent standard deviation. (c) Top panel: Western blot showing levels of SPARC in 30 μg of lysate from RWPE-1, NT2, Cr2, Cr16, and Cr21. SPARC is detectable in RWPE-1 and NT2 but not in the CHD1 KO clones. Bottom panel: loading control. (d) Plot depicting levels of secreted MMP2 protein as detected by ELISA. High levels of MMP2 are secreted by RWPE- 1 compared to the CHD1 KO lines ( ** p < 0.01). NT2 cell lines secreted higher levels of MMP2 than RWPE-1 cell lines ( * p < 0.05). Error bars represent standard deviation. (e) Western blot showing levels of TNC (top panel), VTN (middle panel) and GAPDH (lower panel) in the five cell lines. Cr2 and Cr21 express low levels of TNC compared to RWPE-1 and NT2, while Cr16 expresses TNC levels similar to the parental lines. Cr2 and Cr21 express lower levels of VTN than RWPE-1, while Cr16 expresses similar levels. Bar graphs below show levels of TNC (left) and VTN (right) relative to GAPDH in the five lines. CHD1, chromodomain helicase DNA binding protein 1; ECM, extracellular matrix; ELISA, enzyme-linked immunosorbent assay; FN1, fibronectin; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; ITGA2/4, integrin subunit alpha 2/4; KO, knockout; LAMB3, laminin subunit beta-3 precursor; MMP2/12, matrix metalloproteinase 2/12; NT2, non-target cells; SPARC, secreted protein acidic and rich in cysteine; TNC, tenascin; VTN, vitronectin.

Article Snippet: Primary antibodies used included glyceraldehyde 3-phosphate dehydrogenase (GAPDH) (Santa Cruz Biotechnology Inc., 25778), CHD1 (Novus Biologicals, NB100-60411), focal adhesion kinase (FAK) (Cell Signaling Technologies or CST, 3285P), phosphorylated extracellular signal regulated kinase (pErk) (CST, 4370S), total Erk (CST, 9102S), phosphorylated protein kinase B (pAKT) (CST, 4060S), total AKT (CST, 72), SPARC (CST, 8725), pMEK 1/2 (mitogen-extracellular signal-regulated kinase 1/2) (CST, 9154), total MEK 1/2 (CST, 9126), tenascin C (CST, 12221), and vitronectin (CST, 60896).

Techniques: Expressing, Enzyme-linked Immunosorbent Assay, Standard Deviation, Western Blot, Clone Assay, Control, Binding Assay, Knock-Out

Journal: Veterinary Research

Article Title: Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) moonlights as an adhesin in Mycoplasma hyorhinis adhesion to epithelial cells as well as a plasminogen receptor mediating extracellular matrix degradation

doi: 10.1186/s13567-021-00952-8

Figure Lengend Snippet: Binding of the rGAPDH protein to different ECM components in ELISA experiment . Microtiters plate was coated with Matrigel, fibronectin, collagen or laminin solution. Various concentrations of rGAPDH or BSA were added and detected by anti-His-tag monoclonal antibody. For detecting the binding to vitronectin, microtiters plate was coated with rGAPDH or BSA. Various concentrations of vitronectin were added and detected by anti-vitronectin monoclonal antibody. * P < 0.05, ** P < 0.01, compared with the negative control (BSA).

Article Snippet: The bound vitronectin was detected by rabbit anti-vitronectin monoclonal antibody (1:2000; Boster, China), followed by goat anti-rabbit IgG (1:10 000).

Techniques: Binding Assay, Enzyme-linked Immunosorbent Assay, Negative Control