Journal: Journal of neurological disorders & stroke

Article Title: Neuroendocrine Targeting of Tissue Plasminogen Activator (t-PA)

doi:

Figure Lengend Snippet: A, Microtiter wells were coated with increasing concentrations of DFP-treated t-PA and binding of 1 nM 125I-CgA was determined at 4 hr as described in Experimental Procedures. B, Microtiter wells were coated with DFP-treated t-PA (25 μg/ml) and binding of 1 nM 125I-CgA was determined at the indicated time points. The wells indicated as containing 0 μg/ml t-PA were postcoated with ovalbumin only. Nonspecific binding (circles) was determined in the presence of 1 μM unlabeled CgA and subtracted from total binding (squares) to obtain specific binding (triangles).

Article Snippet: Binding of CgA to Immobilized t-PA Wells of 96 well flat bottom microtiter plates (No. 3912, Falcon, Becton Dickinson, Oxnard, CA) were coated with DFP-inactivated t-PA at 25 μg/ml (unless otherwise indicated) in 0.1 M NaHCO 3 pH 8.5 for 18 hr at 4°C and then postcoated with 3% ovalbumin (Sigma, St. Louis, MO) in phosphate buffered saline (PBS).

Techniques: Binding Assay

Journal: Journal of neurological disorders & stroke

Article Title: Neuroendocrine Targeting of Tissue Plasminogen Activator (t-PA)

doi:

Figure Lengend Snippet: Microtiter wells were coated with t-PA and the interaction with 125I-CgA (1 nM) was determined as described in Experimental Procedures in either the absence or presence of either unlabeled CgA, ovalbumin, RNase, or transferrin (each at 1 μM) for 120 min at 22°C. Total binding is shown.

Article Snippet: Binding of CgA to Immobilized t-PA Wells of 96 well flat bottom microtiter plates (No. 3912, Falcon, Becton Dickinson, Oxnard, CA) were coated with DFP-inactivated t-PA at 25 μg/ml (unless otherwise indicated) in 0.1 M NaHCO 3 pH 8.5 for 18 hr at 4°C and then postcoated with 3% ovalbumin (Sigma, St. Louis, MO) in phosphate buffered saline (PBS).

Techniques: Binding Assay

Journal: Journal of neurological disorders & stroke

Article Title: Neuroendocrine Targeting of Tissue Plasminogen Activator (t-PA)

doi:

Figure Lengend Snippet: Microtiter wells were coated with t-PA and the interaction with 125I-CgA was determined as described in Experimental Procedures in the presence of the indicated concentrations of anti-t-PA mAb's directed against either the EGF-finger domain (open circles), kringle 2 (closed squares), kringle 1 (open squares) or normal mouse IgG (closed circles). Specific binding is shown.

Article Snippet: Binding of CgA to Immobilized t-PA Wells of 96 well flat bottom microtiter plates (No. 3912, Falcon, Becton Dickinson, Oxnard, CA) were coated with DFP-inactivated t-PA at 25 μg/ml (unless otherwise indicated) in 0.1 M NaHCO 3 pH 8.5 for 18 hr at 4°C and then postcoated with 3% ovalbumin (Sigma, St. Louis, MO) in phosphate buffered saline (PBS).

Techniques: Binding Assay

Journal: Journal of neurological disorders & stroke

Article Title: Neuroendocrine Targeting of Tissue Plasminogen Activator (t-PA)

doi:

Figure Lengend Snippet: Microtiter wells were coated with t-PA and incubated with 125I-CgA in either: 10 mM HEPES, pH 7.4, 1 mM EGTA; 10 mM MES pH 6.4 containing 10 mM Ca+2; or 10 mM MES, pH 5.5 containing 20 mM Ca+2. All buffers contained 100 mM NaCl and 0.1%Tween 80. Incubations were performed in the presence of buffer alone or in the presence of either 1 :M CgA, RNAse, or transferrin as indicated.

Article Snippet: Binding of CgA to Immobilized t-PA Wells of 96 well flat bottom microtiter plates (No. 3912, Falcon, Becton Dickinson, Oxnard, CA) were coated with DFP-inactivated t-PA at 25 μg/ml (unless otherwise indicated) in 0.1 M NaHCO 3 pH 8.5 for 18 hr at 4°C and then postcoated with 3% ovalbumin (Sigma, St. Louis, MO) in phosphate buffered saline (PBS).

Techniques: Incubation

Journal: Journal of neurological disorders & stroke

Article Title: Neuroendocrine Targeting of Tissue Plasminogen Activator (t-PA)

doi:

Figure Lengend Snippet: t-PA was immobilized onto wells of microtiter plates. 125I-CgA (1 nM) was incubated with the plates for 4 hours at 22°C in the presence of increasing concentrations of unlabeled CgA. The inhibition curves obtained were transformed to binding isotherms using the LIGAND program. Isotherms were constructed and replotted as Scatchard plots in 10 mM HEPES pH 7.4, 100 mM NaCl, 1 mM EGTA (panels A and D), 10 mM MES pH 6.4, 100 mM NaCl 10 mM Ca+2 (panels B and E) or 10 mM MES pH 5.5, 100 mM NaCl, 20 mM Ca+2 (panels C and F).

Article Snippet: Binding of CgA to Immobilized t-PA Wells of 96 well flat bottom microtiter plates (No. 3912, Falcon, Becton Dickinson, Oxnard, CA) were coated with DFP-inactivated t-PA at 25 μg/ml (unless otherwise indicated) in 0.1 M NaHCO 3 pH 8.5 for 18 hr at 4°C and then postcoated with 3% ovalbumin (Sigma, St. Louis, MO) in phosphate buffered saline (PBS).

Techniques: Incubation, Inhibition, Transformation Assay, Binding Assay, Construct

Journal: Journal of neurological disorders & stroke

Article Title: Neuroendocrine Targeting of Tissue Plasminogen Activator (t-PA)

doi:

Figure Lengend Snippet: A, Microtiter wells were coated with t-PA and incubated with 125I-CgA in the presence of either buffer, unlabeled CgA or each of fourteen synthetic peptides corresponding to bovine CgA (100 μM). B, 125I-CgA was incubated with the wells in the presence of a peptide corresponding to residues 70-90 of CgA (closed squares), or a control peptide corresponding to residues 18-37 of CgA (open squares). Specific binding is shown.

Article Snippet: Binding of CgA to Immobilized t-PA Wells of 96 well flat bottom microtiter plates (No. 3912, Falcon, Becton Dickinson, Oxnard, CA) were coated with DFP-inactivated t-PA at 25 μg/ml (unless otherwise indicated) in 0.1 M NaHCO 3 pH 8.5 for 18 hr at 4°C and then postcoated with 3% ovalbumin (Sigma, St. Louis, MO) in phosphate buffered saline (PBS).

Techniques: Incubation, Control, Binding Assay