E.coli cells expressing recombinant human interferon-γ was disrupted by sonication anddissolved in 7mol·L<sup>-1</sup> guanidine hydrochloride.The extract obtained was then renaturated by 70 folddilu...E.coli cells expressing recombinant human interferon-γ was disrupted by sonication anddissolved in 7mol·L<sup>-1</sup> guanidine hydrochloride.The extract obtained was then renaturated by 70 folddilution with PBS.HulFN γ was purified by affinity chromatography with monoclonal antibody fromthe renaturated crude feed solution.After washing the column with PBS,the adsorbed HulFN γ waseluted with PBS containing 0.5mol·L<sup>-1</sup> NaCl.The column was regenerated with 2mol·L<sup>-1</sup> GuHClfor reuse.After one step of affinity purification the purity of interferon-γ was over 95%.and thespecific activity of the HulFN-γ reached 1.2×10<sup>7</sup> IU·mg<sup>-1</sup> protein.92.8% of recovery was obtainedin the elution step.Total recovery of HulFN γ activity in the affinity chromatography was 78%.展开更多
Recombinant human interferon a2b(rhIFNa2b)is widely used as an antiviral therapy agent for the treatment of hepatitis B and hepatitis C.The current identification test for rhIFNa2b is complex.In this study,an anti-rhI...Recombinant human interferon a2b(rhIFNa2b)is widely used as an antiviral therapy agent for the treatment of hepatitis B and hepatitis C.The current identification test for rhIFNa2b is complex.In this study,an anti-rhIFNa2b nanobody was discovered and used for the development of a rapid lateral flow strip for the identification of rhIFNa2b.RhIFNa2b was used to immunize an alpaca,which established a phage nanobody library.After five steps of enrichment,the nanobody I22,which specifically bound rhIFNa2b,was isolated and inserted into the prokaryotic expression vector pET28a.After subsequent purification,the physicochemical properties of the nanobody were determined.A semiquantitative detection and rapid identification assay of rhIFNa2b was developed using this novel nanobody.To develop a rapid test,the nanobody I22 was coupled with a colloidal gold to produce lateral-flow test strips.The developed rhIFNa2b detection assay had a limit of detection of 1 mg/mL.The isolation of I22 and successful construction of a lateral-flow immunochromatographic test strip demonstrated the feasibility of performing ligand-binding assays on a lateral-flow test strip using recombinant protein products.The principle of this novel assay is generally applicable for the rapid testing of other commercial products,with a great potential for routine use in detecting counterfeit recombinant protein products.展开更多
A new way for the synthesis of human interferon—α_A monoclonal antibody (IFN-α_A-McAb) bound to silica gel packing material in high-performance affinity chromatography (HPAFC) has been developed. The high coupling ...A new way for the synthesis of human interferon—α_A monoclonal antibody (IFN-α_A-McAb) bound to silica gel packing material in high-performance affinity chromatography (HPAFC) has been developed. The high coupling efficiency and specific activity of IFN—α_A-McAb can be obtained by activated diol-silica gel with activating agent. After purification using this packing material in HPAFC, the specific activity of recombinant human interferon-α_A (rIFN-α_A) rose up to 1.03×10~7IU/mg protein and the purification efficiency is appoximately 100 times.展开更多
AIM: To evaluate the antifibrotic effect of different doses of recombinant human Gamma-Interferon (IFN-gamma) in two rat models of hepatic fibrosis, and to observe its effect on moderate chronic hepatitis B virus fibr...AIM: To evaluate the antifibrotic effect of different doses of recombinant human Gamma-Interferon (IFN-gamma) in two rat models of hepatic fibrosis, and to observe its effect on moderate chronic hepatitis B virus fibrosis. METHODS: Hepatic fibrosis was successfully induced in 150 and 196 rats by subcutaneous injection of carbon tetrachloride (CCl4) and intraperitoneal injection of dimethylnitrosamine (DMN), respectively. Each of the two model groups was divided into: (1) fibrotic model group; (2) colchicine treatment group (0.1 mg/kg/day, gastrogavage for 8 weeks); (3) high-dose IFN-gamma group (15 MU/kg per day, i.m. for 8 weeks); (4) medium-dose IFN-gamma group (5 MU/kg daily, i.m. for 8 weeks); and (5) Y low-dose IFN-gamma group (1.67 MU/kg daily, i.m. for 8 weeks). Another group of 10 rats without any treatment was used as normal controls. At the end of the experiment, semi-quantitative histopathological scores of inflammation and fibrosis, liver alpha smooth muscle actin (alpha-SMA) expression level, liver hydroxyl proline content and serum hyaluronic acid levels were compared. And 47 medium chronic hepatitis B viral fibrosis patients were studied. They were given IFN-gamma treatment, 100 MU/day i.m. for the first three months and 100 MU qod i.m. for the next six months. Semi-quantitative pathological scores of inflammation and fibrosis and serum hepatic fibrosis indices were compared within the 9 months. RESULTS: In animal experiment, the pathological fibrosis scores and liver hydroxyl proline content were found to be significantly lower in rats treated with different doses of IFN-gamma as compared with rats in fibrotic model group induced by either CCl4 or DMN, in a dose-dependent manner. For CCl4-induced model, pathological fibrosis scores in high, medium and low doses IFN-gamma groups were 5.10 +/- 2.88, 7.70 +/- 3.53 and 8.00 +/- 3.30, respectively, but the score was 14.60 +/- 7.82 in fibrotic model group. Hydroxyl proline contents were 2.83 +/- 1.18, 3.59 +/- 1.22 and 4.80 +/- 1.62, in the three IFN-gamma groups, and 10.01 +/- 3.23 in fibrotic model group. The difference was statistically significant (P【0.01). Similar results were found in DMN-induced model. Pathological fibrosis scores were 6.30 +/- 0.48, 8.10 +/- 2.72 and 8.30 +/- 2.58, in high, medium and low doses IFN-gamma groups, and 12.60 +/- 3.57 in fibrotic model group. Hydroxyl proline contents were 2.72 +/- 0.58, 3.14 +/- 0.71 and 3.62 +/- 1.02, in the three IFN-gamma groups, and 12.79 +/- 1.54 in fibrotic model group. The difference was statistically significant (P【0.01).Serum hepatic fibrosis indices decreased significantly in the 47 patients after IFN-gamma treatment (HA: 433.38 +/- 373.00 vs 281.57 +/- 220.48; LN: 161.22 +/- 41.02 vs 146 +/- 35 +/- 44. 67; PC III: 192.59 +/- 89.95 vs 156.98 +/- 49.22; C-I: 156.30 +/- 44.01 vs 139.14 +/- 34.47) and the differences between the four indices were significant (P 【0.05). Thirty-three patients received two liver biopsies, one before and one after IFN-gamma treatment. In thirty of 33 patients IFN-gamma had better effects according to semi-quantitative pathological scores (8.40 +/- 5.83 vs 5.30 +/- 4.05, P【0.05). CONCLUSION: All the three doses of IFN-gamma are effective in treating rat liver fibrosis induced by either CCl4 or DMN, the higher the dose, the better the effect. And IFN-gamma is effective for patients with moderate chronic hepatitis B viral fibrosis.展开更多
目的观察重组人干扰素α2b(recombinant human interferon-α2b,rhIFN-α2b)和拉米夫定序贯治疗用于乙肝免疫耐受期患儿的效果。方法选择西安医学院第三附属医院2020年4月至2023年4月收治的96例乙肝免疫耐受期患儿为研究对象,按随机数...目的观察重组人干扰素α2b(recombinant human interferon-α2b,rhIFN-α2b)和拉米夫定序贯治疗用于乙肝免疫耐受期患儿的效果。方法选择西安医学院第三附属医院2020年4月至2023年4月收治的96例乙肝免疫耐受期患儿为研究对象,按随机数字法分为研究组与对照组各48例。研究组男29例,女19例;年龄(7.54±1.36)岁;病程(2.25±0.54)年;有乙肝家族史28例。对照组男26例,女22例;年龄(7.63±1.41)岁;病程(2.44±0.51)年;有乙肝家族史26例。对照组口服拉米夫定片,0.1 g/次,1次/d,治疗24周。研究组采用rhIFN-α2b和拉米夫定序贯治疗:前4周单用rhIFN-α2b肌内或皮下注射,5 mIU/(m^(2)·次),1次/2 d,4周后加用拉米夫定,0.1 g/次,1次/d,持续治疗8周后停用rhIFN-α2b仅用拉米夫定,继续治疗12周。比较两组疗效;治疗4周、12周、24周后,记录两组乙肝病毒E抗原(hepatitis B virus E antigen,HBeAg)、乙肝病毒脱氧核糖核酸(hepatitis B virus DNA,HBV-DNA)转阴率和乙肝病毒E抗体(hepatitis B virus E antibody,HBeAb)转换率;治疗前及治疗24周后,比较两组肝功能[谷丙转氨酶(alanine aminotransferase,ALT)、谷草转氨酶(aspertate aminotransferase,AST)、总胆红素(total bilirubin,TBil)]及肝纤维化指标[透明质酸(hyaluronic acid,HA)、层黏蛋白(laminin,LN)、Ⅲ型前胶原肽(type Ⅲ procollagen peptide,PⅢP)、Ⅳ型胶原(type Ⅳ collagen,CⅣ)];记录两组不良反应发生情况。采用t检验、χ^(2)检验进行统计学分析。结果研究组总有效率高于对照组[77.08%(37/48)比52.08%(25/48)](P<0.05)。治疗4周及12周后,研究组与对照组HBeAg、HBV-DNA转阴率比较差异均无统计学意义(均P>0.05),治疗24周后,研究组HBeAg、HBV-DNA转阴率均高于对照组(均P<0.05);两组治疗后不同时间点HBeAb转换率比较差异均无统计学意义(均P>0.05)。治疗24周后,两组ALT、AST、TBil均降低,且研究组[(39.16±7.51)U/L、(41.92±8.26)U/L、(15.13±4.29)μmol/L]均低于对照组[(47.38±8.02)U/L、(52.36±8.73)U/L、(18.67±4.86)μmol/L](均P<0.05)。治疗24周后,两组HA、LN、PⅢP、CⅣ均降低,且研究组[(113.57±30.16)μg/L、(96.41±29.05)μg/L、(78.14±20.96)μg/L、(90.26±26.31)μg/L]均低于对照组[(208.34±64.72)μg/L、(124.27±32.19)μg/L、(104.37±22.48)μg/L、(143.75±33.49)μg/L](均P<0.05)。两组不良反应发生率比较差异无统计学意义(P>0.05)。结论rhIFN-α2b和拉米夫定序贯治疗在乙肝免疫耐受期患儿中疗效确切,能有效减少病毒含量,阻止肝功能恶化及肝纤维化,且安全性较高。展开更多
目的研究宫颈高危型HPV感染应用重组人干扰素α2b阴道泡腾胶囊治疗的作用。方法选取100例宫颈高危型HPV感染患者,根据随机数字表法分为研究组与一般组,各50例。一般组定期随访不做任何处理,研究组采用重组人干扰素α2b阴道泡腾胶囊治疗...目的研究宫颈高危型HPV感染应用重组人干扰素α2b阴道泡腾胶囊治疗的作用。方法选取100例宫颈高危型HPV感染患者,根据随机数字表法分为研究组与一般组,各50例。一般组定期随访不做任何处理,研究组采用重组人干扰素α2b阴道泡腾胶囊治疗。比较两组转阴情况、治疗前后生活质量评分。结果研究组转阴率高于(62.00 vs 22.00)%一般组(P<0.05)。治疗后,研究组生活质量评分(75.98±10.52 vs 85.98±11.03)分高于一般组(P<0.05)。结论宫颈高危型HPV感染患者实施重组人干扰素α2b阴道泡腾胶囊治疗,可有效提高转阴率,改善患者生活质量,值得临床推广。展开更多
文摘E.coli cells expressing recombinant human interferon-γ was disrupted by sonication anddissolved in 7mol·L<sup>-1</sup> guanidine hydrochloride.The extract obtained was then renaturated by 70 folddilution with PBS.HulFN γ was purified by affinity chromatography with monoclonal antibody fromthe renaturated crude feed solution.After washing the column with PBS,the adsorbed HulFN γ waseluted with PBS containing 0.5mol·L<sup>-1</sup> NaCl.The column was regenerated with 2mol·L<sup>-1</sup> GuHClfor reuse.After one step of affinity purification the purity of interferon-γ was over 95%.and thespecific activity of the HulFN-γ reached 1.2×10<sup>7</sup> IU·mg<sup>-1</sup> protein.92.8% of recovery was obtainedin the elution step.Total recovery of HulFN γ activity in the affinity chromatography was 78%.
基金support was provided by the National Science and Technology Major Project(Grant No.:2015ZX09501008)。
文摘Recombinant human interferon a2b(rhIFNa2b)is widely used as an antiviral therapy agent for the treatment of hepatitis B and hepatitis C.The current identification test for rhIFNa2b is complex.In this study,an anti-rhIFNa2b nanobody was discovered and used for the development of a rapid lateral flow strip for the identification of rhIFNa2b.RhIFNa2b was used to immunize an alpaca,which established a phage nanobody library.After five steps of enrichment,the nanobody I22,which specifically bound rhIFNa2b,was isolated and inserted into the prokaryotic expression vector pET28a.After subsequent purification,the physicochemical properties of the nanobody were determined.A semiquantitative detection and rapid identification assay of rhIFNa2b was developed using this novel nanobody.To develop a rapid test,the nanobody I22 was coupled with a colloidal gold to produce lateral-flow test strips.The developed rhIFNa2b detection assay had a limit of detection of 1 mg/mL.The isolation of I22 and successful construction of a lateral-flow immunochromatographic test strip demonstrated the feasibility of performing ligand-binding assays on a lateral-flow test strip using recombinant protein products.The principle of this novel assay is generally applicable for the rapid testing of other commercial products,with a great potential for routine use in detecting counterfeit recombinant protein products.
文摘A new way for the synthesis of human interferon—α_A monoclonal antibody (IFN-α_A-McAb) bound to silica gel packing material in high-performance affinity chromatography (HPAFC) has been developed. The high coupling efficiency and specific activity of IFN—α_A-McAb can be obtained by activated diol-silica gel with activating agent. After purification using this packing material in HPAFC, the specific activity of recombinant human interferon-α_A (rIFN-α_A) rose up to 1.03×10~7IU/mg protein and the purification efficiency is appoximately 100 times.
文摘AIM: To evaluate the antifibrotic effect of different doses of recombinant human Gamma-Interferon (IFN-gamma) in two rat models of hepatic fibrosis, and to observe its effect on moderate chronic hepatitis B virus fibrosis. METHODS: Hepatic fibrosis was successfully induced in 150 and 196 rats by subcutaneous injection of carbon tetrachloride (CCl4) and intraperitoneal injection of dimethylnitrosamine (DMN), respectively. Each of the two model groups was divided into: (1) fibrotic model group; (2) colchicine treatment group (0.1 mg/kg/day, gastrogavage for 8 weeks); (3) high-dose IFN-gamma group (15 MU/kg per day, i.m. for 8 weeks); (4) medium-dose IFN-gamma group (5 MU/kg daily, i.m. for 8 weeks); and (5) Y low-dose IFN-gamma group (1.67 MU/kg daily, i.m. for 8 weeks). Another group of 10 rats without any treatment was used as normal controls. At the end of the experiment, semi-quantitative histopathological scores of inflammation and fibrosis, liver alpha smooth muscle actin (alpha-SMA) expression level, liver hydroxyl proline content and serum hyaluronic acid levels were compared. And 47 medium chronic hepatitis B viral fibrosis patients were studied. They were given IFN-gamma treatment, 100 MU/day i.m. for the first three months and 100 MU qod i.m. for the next six months. Semi-quantitative pathological scores of inflammation and fibrosis and serum hepatic fibrosis indices were compared within the 9 months. RESULTS: In animal experiment, the pathological fibrosis scores and liver hydroxyl proline content were found to be significantly lower in rats treated with different doses of IFN-gamma as compared with rats in fibrotic model group induced by either CCl4 or DMN, in a dose-dependent manner. For CCl4-induced model, pathological fibrosis scores in high, medium and low doses IFN-gamma groups were 5.10 +/- 2.88, 7.70 +/- 3.53 and 8.00 +/- 3.30, respectively, but the score was 14.60 +/- 7.82 in fibrotic model group. Hydroxyl proline contents were 2.83 +/- 1.18, 3.59 +/- 1.22 and 4.80 +/- 1.62, in the three IFN-gamma groups, and 10.01 +/- 3.23 in fibrotic model group. The difference was statistically significant (P【0.01). Similar results were found in DMN-induced model. Pathological fibrosis scores were 6.30 +/- 0.48, 8.10 +/- 2.72 and 8.30 +/- 2.58, in high, medium and low doses IFN-gamma groups, and 12.60 +/- 3.57 in fibrotic model group. Hydroxyl proline contents were 2.72 +/- 0.58, 3.14 +/- 0.71 and 3.62 +/- 1.02, in the three IFN-gamma groups, and 12.79 +/- 1.54 in fibrotic model group. The difference was statistically significant (P【0.01).Serum hepatic fibrosis indices decreased significantly in the 47 patients after IFN-gamma treatment (HA: 433.38 +/- 373.00 vs 281.57 +/- 220.48; LN: 161.22 +/- 41.02 vs 146 +/- 35 +/- 44. 67; PC III: 192.59 +/- 89.95 vs 156.98 +/- 49.22; C-I: 156.30 +/- 44.01 vs 139.14 +/- 34.47) and the differences between the four indices were significant (P 【0.05). Thirty-three patients received two liver biopsies, one before and one after IFN-gamma treatment. In thirty of 33 patients IFN-gamma had better effects according to semi-quantitative pathological scores (8.40 +/- 5.83 vs 5.30 +/- 4.05, P【0.05). CONCLUSION: All the three doses of IFN-gamma are effective in treating rat liver fibrosis induced by either CCl4 or DMN, the higher the dose, the better the effect. And IFN-gamma is effective for patients with moderate chronic hepatitis B viral fibrosis.
文摘目的观察重组人干扰素α2b(recombinant human interferon-α2b,rhIFN-α2b)和拉米夫定序贯治疗用于乙肝免疫耐受期患儿的效果。方法选择西安医学院第三附属医院2020年4月至2023年4月收治的96例乙肝免疫耐受期患儿为研究对象,按随机数字法分为研究组与对照组各48例。研究组男29例,女19例;年龄(7.54±1.36)岁;病程(2.25±0.54)年;有乙肝家族史28例。对照组男26例,女22例;年龄(7.63±1.41)岁;病程(2.44±0.51)年;有乙肝家族史26例。对照组口服拉米夫定片,0.1 g/次,1次/d,治疗24周。研究组采用rhIFN-α2b和拉米夫定序贯治疗:前4周单用rhIFN-α2b肌内或皮下注射,5 mIU/(m^(2)·次),1次/2 d,4周后加用拉米夫定,0.1 g/次,1次/d,持续治疗8周后停用rhIFN-α2b仅用拉米夫定,继续治疗12周。比较两组疗效;治疗4周、12周、24周后,记录两组乙肝病毒E抗原(hepatitis B virus E antigen,HBeAg)、乙肝病毒脱氧核糖核酸(hepatitis B virus DNA,HBV-DNA)转阴率和乙肝病毒E抗体(hepatitis B virus E antibody,HBeAb)转换率;治疗前及治疗24周后,比较两组肝功能[谷丙转氨酶(alanine aminotransferase,ALT)、谷草转氨酶(aspertate aminotransferase,AST)、总胆红素(total bilirubin,TBil)]及肝纤维化指标[透明质酸(hyaluronic acid,HA)、层黏蛋白(laminin,LN)、Ⅲ型前胶原肽(type Ⅲ procollagen peptide,PⅢP)、Ⅳ型胶原(type Ⅳ collagen,CⅣ)];记录两组不良反应发生情况。采用t检验、χ^(2)检验进行统计学分析。结果研究组总有效率高于对照组[77.08%(37/48)比52.08%(25/48)](P<0.05)。治疗4周及12周后,研究组与对照组HBeAg、HBV-DNA转阴率比较差异均无统计学意义(均P>0.05),治疗24周后,研究组HBeAg、HBV-DNA转阴率均高于对照组(均P<0.05);两组治疗后不同时间点HBeAb转换率比较差异均无统计学意义(均P>0.05)。治疗24周后,两组ALT、AST、TBil均降低,且研究组[(39.16±7.51)U/L、(41.92±8.26)U/L、(15.13±4.29)μmol/L]均低于对照组[(47.38±8.02)U/L、(52.36±8.73)U/L、(18.67±4.86)μmol/L](均P<0.05)。治疗24周后,两组HA、LN、PⅢP、CⅣ均降低,且研究组[(113.57±30.16)μg/L、(96.41±29.05)μg/L、(78.14±20.96)μg/L、(90.26±26.31)μg/L]均低于对照组[(208.34±64.72)μg/L、(124.27±32.19)μg/L、(104.37±22.48)μg/L、(143.75±33.49)μg/L](均P<0.05)。两组不良反应发生率比较差异无统计学意义(P>0.05)。结论rhIFN-α2b和拉米夫定序贯治疗在乙肝免疫耐受期患儿中疗效确切,能有效减少病毒含量,阻止肝功能恶化及肝纤维化,且安全性较高。
文摘目的研究宫颈高危型HPV感染应用重组人干扰素α2b阴道泡腾胶囊治疗的作用。方法选取100例宫颈高危型HPV感染患者,根据随机数字表法分为研究组与一般组,各50例。一般组定期随访不做任何处理,研究组采用重组人干扰素α2b阴道泡腾胶囊治疗。比较两组转阴情况、治疗前后生活质量评分。结果研究组转阴率高于(62.00 vs 22.00)%一般组(P<0.05)。治疗后,研究组生活质量评分(75.98±10.52 vs 85.98±11.03)分高于一般组(P<0.05)。结论宫颈高危型HPV感染患者实施重组人干扰素α2b阴道泡腾胶囊治疗,可有效提高转阴率,改善患者生活质量,值得临床推广。
