The only difference of primary structure between single-chain prourokinase (pro-UK or scu-PA) and two-chain urokinase (UK or tcu-PA) is the cleavage of a single peptide bond (Lys158-Ile159) and transform scu-PA into i...The only difference of primary structure between single-chain prourokinase (pro-UK or scu-PA) and two-chain urokinase (UK or tcu-PA) is the cleavage of a single peptide bond (Lys158-Ile159) and transform scu-PA into its active two-chain form. A 13-peptide (Thr-Leu-Arg-Pro-Arg-Phe-Lys-Ile-Ile-Gly- Gly-Glu-Cys), which spans the cleavage peptide bond, was synthesized and linked to KLH (Keyhole limpet hemocyanin). The Balb/c mice were immunized by the conjugated protein with proper adjuvant. According to the Kohler and Milstein’s methods, a hybridoma cell line G7 secreting monoclonal antibody specific for scu-PA was obtained. The anti-scu-PA McAb, purified from the supernatant of porous microcarrier hybridoma cell culture, was conjugated to CNBr-activated Sepharose 4B to prepare an immuno-affinity chromatography column. The u-PA was purified only by this affinity column from the supernatant of cultivating the u-PA-producing recombinant CHO cell, the u-PA recovery ratio is 90.4%, the purification factor was about 50, with the specific activity of 1.2×10 5IU/mg, the scu-PA ratio in the u-PA product was 96.3%. Compared to immuno-affinity chromatography, the 3-step process for purifying u-PA (cation-exchange co-lumn, gel filtration column and benzamidine affinity column)has a u-PA recovery ratio of about 65%, with a specific activity of 1.0×10 5IU/mg, and an scu-PA ratio of about 90%. These results showed that immuno-affinity chromatography is simple to recover u-PA and effective to separate scu-PA from tcu-PA.展开更多
PURPOSE: To study the results of intravitreal tissue plasminogen activator (tPA) and expansile gas injection for submacular haemorrhage in Thai patients. Methods: The medical records of Thai patients who presented wit...PURPOSE: To study the results of intravitreal tissue plasminogen activator (tPA) and expansile gas injection for submacular haemorrhage in Thai patients. Methods: The medical records of Thai patients who presented with submacular haemorrhage between January 1998 and December 2002 were reviewed. The inclusion criteria were acute onset of bleeding (<1 month), treatment with intravitreal injection of tPA solution (50-100 μg in 0.1ml) and expansile gas (0.3-0.4ml of 100%perfluoropropane or sulphur hexafluoride), and at least 6 months of follow-up. Our main outcome measures were best final postoperative visual acuity and surgical complications. Results: A total of 19 eyes of 19 patients completed the inclusion criteria with a mean duration of 13.1 days. The causes of haemorrhage were age-related macular degeneration in 15 eyes (78.9%), idiopathic choroidal neovascularization in two eyes (10.5%), and traumatic, and valsalva retinopathy in one eye each (5.2%). After a mean follow-up of 13 months (range 6-39 months), postoperative visual acuity improved two lines or greater in 12 eyes (63.2%), stabilized in six eyes (31.6%) and worsened in one (5.2%). The final visual acuity measured 20/63 or better in 10 eyes (52.6%). The surgical complications were breakthrough vitreous haemorrhage (three eyes) and cataracts (three eyes), and two had retinal detachments. Conclusion: The treatment of submacular haemorrhage with intravitreal injection of tPA and expansile gas improved visual acuity in more than half of the patients. In all, 10 in 19 eyes demonstrated final visual acuity at a functional level.展开更多
文摘The only difference of primary structure between single-chain prourokinase (pro-UK or scu-PA) and two-chain urokinase (UK or tcu-PA) is the cleavage of a single peptide bond (Lys158-Ile159) and transform scu-PA into its active two-chain form. A 13-peptide (Thr-Leu-Arg-Pro-Arg-Phe-Lys-Ile-Ile-Gly- Gly-Glu-Cys), which spans the cleavage peptide bond, was synthesized and linked to KLH (Keyhole limpet hemocyanin). The Balb/c mice were immunized by the conjugated protein with proper adjuvant. According to the Kohler and Milstein’s methods, a hybridoma cell line G7 secreting monoclonal antibody specific for scu-PA was obtained. The anti-scu-PA McAb, purified from the supernatant of porous microcarrier hybridoma cell culture, was conjugated to CNBr-activated Sepharose 4B to prepare an immuno-affinity chromatography column. The u-PA was purified only by this affinity column from the supernatant of cultivating the u-PA-producing recombinant CHO cell, the u-PA recovery ratio is 90.4%, the purification factor was about 50, with the specific activity of 1.2×10 5IU/mg, the scu-PA ratio in the u-PA product was 96.3%. Compared to immuno-affinity chromatography, the 3-step process for purifying u-PA (cation-exchange co-lumn, gel filtration column and benzamidine affinity column)has a u-PA recovery ratio of about 65%, with a specific activity of 1.0×10 5IU/mg, and an scu-PA ratio of about 90%. These results showed that immuno-affinity chromatography is simple to recover u-PA and effective to separate scu-PA from tcu-PA.
文摘PURPOSE: To study the results of intravitreal tissue plasminogen activator (tPA) and expansile gas injection for submacular haemorrhage in Thai patients. Methods: The medical records of Thai patients who presented with submacular haemorrhage between January 1998 and December 2002 were reviewed. The inclusion criteria were acute onset of bleeding (<1 month), treatment with intravitreal injection of tPA solution (50-100 μg in 0.1ml) and expansile gas (0.3-0.4ml of 100%perfluoropropane or sulphur hexafluoride), and at least 6 months of follow-up. Our main outcome measures were best final postoperative visual acuity and surgical complications. Results: A total of 19 eyes of 19 patients completed the inclusion criteria with a mean duration of 13.1 days. The causes of haemorrhage were age-related macular degeneration in 15 eyes (78.9%), idiopathic choroidal neovascularization in two eyes (10.5%), and traumatic, and valsalva retinopathy in one eye each (5.2%). After a mean follow-up of 13 months (range 6-39 months), postoperative visual acuity improved two lines or greater in 12 eyes (63.2%), stabilized in six eyes (31.6%) and worsened in one (5.2%). The final visual acuity measured 20/63 or better in 10 eyes (52.6%). The surgical complications were breakthrough vitreous haemorrhage (three eyes) and cataracts (three eyes), and two had retinal detachments. Conclusion: The treatment of submacular haemorrhage with intravitreal injection of tPA and expansile gas improved visual acuity in more than half of the patients. In all, 10 in 19 eyes demonstrated final visual acuity at a functional level.
