Characterization of indium-111 labeled recombinant tissue plasminogen activator for the imaging of thrombi

Abstract

The in vitro functional properties of recombinant tissue plasminogen activator (rt-PA), its biodistribution in mice, and its pharmacokinetics and clot localization properties in dogs have been investigated after labeling rt-PA with 111In. The rt-PA was coupled with the bicyclic anhydride of DTPA using standard methodology. Amidolytic and fibrinolytic assays showed retention of protein activity when rt-PA was conjugated with an average of one DTPA group or less per molecule. Size exclusion HPLC showed each preparation to be radiochemically pure with 111In bound exclusively to the attached DTPA groups. Biodistribution in mice showed major accumulation of activity in the liver and kidneys. After administration of 0.5-1.0 mg of the labeled protein to dogs, blood activity decreased with a half time of approximately 5 min in agreement with previous reports of rapid blood clearance. Largely because of decreased blood levels, clot: blood ratios of labeled protein increased rapidly, in one study reaching 6.3 after 31 min, and satisfactory images of fibrin thrombi were obtained. The rt-PA may be labeled with 111In without destroying the ability of the protein to localize in clot and images of forming clot can be obtained with this agent within 1 h after administration.

Type VI Collagen Anchors Endothelial Basement Membranes by Interacting with Type IV Collagen

“…Affinity Blot-Purified type VI collagen tetramers and globular do-mains were biotinylated according to the method of Hnatowich et al (22) with minor modifications. To modify the primary amino groups of type VI collagen and its fragments, the purified samples were dissolved into 0.5 M Tris-HCl, pH 8.0, buffer containing 8 M urea, 0.2 M NaCl, and 5 mM EDTA at a final protein concentration of 0.5 mg/ml and dialyzed against 50 mM sodium bicarbonate, pH 8.4, containing 0.1 M NaCl.…”

A transition metal complex (Venus flytrap cluster) for radioimmunodetection and radioimmunotherapy.

“…These conjugates form stable radiopharmaceuticals with 1"'In that have been used for radioimmunodiagnosis of human tumors (6)(7)(8), although there is some concern about their uptake by normal tissues and possible metal release in vivo. Conjugates of this type, however, form less stable radiopharmaceuticals with 90Y and 67Cu (9,10), which can be used for radioimmunotherapy. There is therefore considerable interest in the development of new bifunctional reagents that will form more stable chelate complexes with these and other clinically relevant radionuclides.…”

An SECM Detection Scheme with Improved Sensitivity and Lateral Resolution: Detection of Galactosidase Activity with Signal Amplification by Glucose Dehydrogenase

“…[7] Apo-GDH (PQQ-dependent, from Escherichia coli, EC 1.1.99.17) was biotinylated, and then a Lowry protein assay was performed to determine the concentration of the enzyme. [10] Biotinylated GDH was treated similarly. [7] To reconstitute the holo-GDH from apo-GDH, PQQ (0.5 mg mL À1 , 50 mL) and CaCl 2 (0.1m, 10 mL) were added to a suspension of the beads (40 mL), and the mixture was incubated at room temperature for 30 min.…”