Time filter

Source Type

Qian F.,Shanghai University | Li M.,Shanghai University | Chen Y.,Lanzhou Institute of Biological Products Co. | Jiang L.,Lanzhou Institute of Biological Products Co. | Xu H.,Shanghai University
Human Vaccines and Immunotherapeutics | Year: 2016

Plasmodium falciparum surface protein 25 (Pfs25) is a hard-to-express and hard-to-solubilize protein in Escherichia coli. To overcome this problem, the phase 1 flagellin of Salmonella enterica serovar Typhimurium (FliC) was used as a fusion partner for Pfs25. The fusion expression of Pfs25 with FliC greatly enhanced the expression level and solubility of Pfs25 in E. coli BL21(DE3). The Ni-purified fusion protein of FliC-Pfs25 was recognized by two anti-Pfs25 monoclonal antibodies. By comparison, it was shown that the Pfs25 within FliC-Pfs25 contained epitopes similar or identical to those on Pichia pastoris-produced Pfs25. The data obtained from this study demonstrated that the fusion with Salmonella flagellin greatly improved the expression of Pfs25 in E. coli. © 2016 Taylor & Francis Group, LLC.


He X.,Lanzhou Institute of Biological Products Co.
Chinese Journal of Biologicals | Year: 2012

Objective: To analyze the property of active pharmaceutical ingredient (API) of botulinum toxin type A (BTXA) for injection, as well as the specific activities of the preparations manufactured in 2009-2011. Methods: Various components were separated from the harvested BTXA by anion exchange chromatography. The properties of BTXA complex and its components were analyzed by hemagglutination test, HPLC, SDS-PAGE, isoelectric focusing electrophoresis and amino acid sequencing at N-terminus. The data on specific activity of bulks of BTXA manufactured in 2009-2011 were analyzed, based on which the stability of production procedure and the reproducibility of quality were evaluated. Results: The harvested BTXA was dissociated in basic environment. The obtained neurotoxin showed no hemagglutination titer, of which the relative molecular mass was about 150 000. The API of BTXA was an intact monomer, of which the purity was more than 99.5%. The isoelectric points of BTXA complex and neurotoxin were 4.97 and 4.91 respectively, of which the amino acid sequences at N-terminus were basically consistent with those reported in GenBank. The mean specific activity of bulks of BTXA manufactured in 2009-2011 was 3.0 × 107 LD50/mg protein. The load of API in final product of BTXA was about 5 ng per container. Conclusion: The API of BTXA was a specific complex which was dissociated in basic environment to obtain neurotoxin. The specific activities of bulk BTXA manufactured in 2009-2011 were stable, indicating good persistency of quality, which provided a basis for safety of BTXA in clinic.


Fu Y.-X.,Lanzhou Institute of Biological Products Co. | Ma Q.-H.,Lanzhou Institute of Biological Products Co. | Feng X.,Lanzhou Institute of Biological Products Co. | Luo L.-H.,Lanzhou Institute of Biological Products Co. | And 2 more authors.
Chinese Journal of Biologicals | Year: 2014

Objective: To develop a method for determination of residual sodium periodate (NaIO4) content in meningococcal polysaccharide protein conjugate vaccine. Methods: Periodate was reduced to iodide ion with ascorbic acid, then loaded onto IonPacAS11-HC anion exchange column (4 mm x 250 mm) at a loading of 25 μl, eluted with 50 mmol/L sodium hydroxide at a flow rate of 1.5 ml/min, and determined with electrochemical detector, of which the data were recorded and analyzed by using Chromeleon chromatography workstation. The developed method was verified for specificity, accuracy and reproducibility, determined for minimum detection and quantitative detection limits, and applied preliminarily. Results: Periodate was completely reduced to iodide ion when the proportion of ascorbic acid to sodium periodate, at equal concentrations, was 4:1. The iodide ion concentration of control at a range of 10 μg/L-1 mg/L showed good liner relationship to chromatographic peak area, with a r value of more than 0.999. The recovery rate, RSD, minimum detection limit and minimum quantitative detection limit of the developed method were 91.66%-110.20%, 0.2%-2.3%, 1 μg/L [with a signal-to-noise ratio (SNR) of 3:1] and 5 μg/L (with a SNR of 10:1) respectively. Each of two batches of meningococcal polysaccharide protein conjugate vaccine of groups A, C, Y and W135 were determined by the developed method, and no sodium periodate was detected. Conclusion: The developed method was simple, sensitive, rapid, with little interference and good reproducibility, which was suitable for the quality control of vaccine during production.


Duan L.-J.,Lanzhou Institute of Biological Products Co. | Yang J.-J.,Lanzhou Institute of Biological Products Co. | Zhang J.,Lanzhou Institute of Biological Products Co.
Chinese Journal of Biologicals | Year: 2014

Objective: To purify group A botulinum antitoxin by chromatography so as to increase the purity of immunoglobulin F(ab')2. Methods: Group A botulinum antitoxin was purified by anion-exchange chromatography. The pH value and conductivity of buffer were optimized by Design of Experiments (DoE). Results: The low molecular impurity content in flow through buffer was influenced significantly by conductivity, which decreased with the decreasing conductivity. However, pH value with the range of DoE showed little effect on the content. The aggregate/polymer content was high at low pH values and high conductivity, while was low at high pH values and low conductivity. The purity of F(ab')2 was mainly influenced by conductivity, which decreased with the increasing conductivity, and was more than 90% when the conductivity was within the DoE range (2 ∼ 20 ms/cm). Conclusion: Anion-exchange chromatography decreased the aggregate/polymer and partial low molecular impurity contents in group A botulinum antitoxin and increased the purity of F(ab')2.


An C.,Lanzhou Institute of Biological Products Co. | Mao X.-Y.,Lanzhou Institute of Biological Products Co.
Chinese Journal of Biologicals | Year: 2014

With the increasing needs for therapeutic monoclonal antibodies in market, the study in this field has focused on how to obtain a high yield and stable mammalian cell strain, which is mainly challenged by shortening the development process so as to obtain high yield and stable expression cell line. In the whole development process of therapeutic monoclonal antibodies, the design and optimization of mammalian cell expression vector as the first step play a particularly important role. This paper reviews the strategy for design and optimization of expression vector for therapeutic monoclonal antibody, including action element and mode at gene transcription level, label of gene amplification and screening, position effect as well as optimization of integration site.


Chang Y.-L.,Lanzhou Institute of Biological Products Co.
Chinese Journal of Biologicals | Year: 2012

Objective: To determine the F1 and rV antigen contents in plague component vaccine by double antibody sandwich ELISA. Methods: The antigens in bulk of plague component vaccine were identified with monoclonal antibodies (McAbs) against F1 rV antigens, and determined quantitatively by double antibody sandwich ELISA. The completeness of antigen adsorbed onto aluminium hydroxide adjuvant was verified. The final product of vaccine was stored at 4°C for 18 months, from which samples were taken 1 and 18 months after storage respectively and determined for F1 and rV antigen contents by the developed double antibody sandwich ELISA method to analyze the stability of antigen. The three-step purification procedure for three batches of bulk rV antigen of Yersiria pestis were verified. Results: The F1 and rV antigens in bulk of plague component vaccine showed good reactogenicity. The F1 and rV antigen contents in four batches of bulks were determined by the developed method, of which the results showed an error of not more than 20% with those determined by Lowry method. No F1 or rV antigen was detected in the centrifuge supernatant of four batches vaccine, indicating that the antigens were completely adsorbed by aluminium hydroxide adjuvant. The F1 and rV antigen contents in vaccine were stable after storage at 4°C for 18 months. After three batches of bulk were purified by anion exchange, hydrophobic and gel filtration chromatography, both the purity and content of rV antigen increased. Conclusion: The developed double antibody sandwich ELISA method may be used for the quantitative determination of F1 and rV antigens in plague component vaccine.


Wang J.F.,Lanzhou Institute of Biological Products Ltd | Mao X.Y.,Lanzhou Institute of Biological Products Ltd | Zhao C.,Lanzhou Institute of Biological Products Ltd
Molecular Biology Reports | Year: 2014

The experiment were performed to investigate the poisoning-related proteins and main pathological changes after mouse suffered from injection of botulinum toxin serotype E. Dose of 0.75 LD50 botulinum toxin serotype E per mice were administrated by intraperitoneal injection. Survival mouse were picked as experimental group. The blood were collected from orbital blood and serum sample was separated by centrifugation. The heart, liver, spleen, lung, kidney were fixed in 10 % neutral buffered formalin and then developed paraffin sections. Serum protein components were analyzed by SDS-PAGE gel electrophoresis coupled with 2-DE SDS-PAGE gel electrophoresis. Differentially expressed proteins were analyzed by PDQUest8.0 software and subjected to ion trap mass spectrometry equipped with a high performance liquid chromatography system. The observation of pathological section showed that heart, liver, spleen, lung, kidney exhibited pathological changes in different degree, especially in heart, liver and lung tissues. Heart muscle tissue display serious inflammatory response, heart muscle fiber compulsively expanded and filled with erythrocyte and inflammatory exudates, some heart muscle fiber ruptured, even necrosis; hepatic cell in edge of liver occur apoptosis and some hepatic cell have disintegrated, and even died; pulmonary alveoli broken and partial vein filled with blood. Serum proteins component present a significant changes between control serum and botulism in 24 h by SDS-PAGE gel electrophoresis and 2-DE-SDS-PAGE gel electrophoresis. Twenty differentially expressed protein spots were observed in 2-DE profiles, in which 14 protein spots were undetectable in serum proteome under botulism, 3 protein spots exclusively expressed in state of botulism, 3 protein spots were low-expressed in serum proteome under botulism. Fourteen proteins have been identified among 20 spots elected on two-dimensional electrophoresis gels. Crystal proteins family exclusively expressed in control group serum. Haptoglobin were low-expressed under botulism in serum protein components, however, serum amyloid A only expressed in serum sample under botulism in 24 h, which were verified by Western-blot. Identified proteins involved in energy metabolism, cellular stress response, transcription, body defense and cell proliferation. These findings represent the first report of BoNT-induced changes in serum proteome and histopathology, and reinforce the utility of applying proteomic tools to the study of system-wide biological processes in normal and botulism. © 2014 Springer Science+Business Media Dordrecht.


Chang Y.-L.,Lanzhou Institute of Biological Products Co.
Chinese Journal of Biologicals | Year: 2013

Objective: To develop and verify a double antibody sandwich ELISA diagnostic kit for F1 antigen of Yersinia pestis. Methods: Antisera were prepared by immunizing rabbits with F1 antigen of Y. pestis, subjected to salting out with 50% ammonium sulfate, the purified by Sephacryl S-300 gle column chromatography and used as the coating antibody. The murine ascites containing monoclonal antibody (McAb) against F1 antigen was prepared, purified, labled with HRP and used as the enzyme-labeled antibody. Three batches of double antibody sandwich ELISA kits were prepared, and verified for precsion, sensitivity, linear range, specificity, reproducibility, accuracy and stability by intra- and inter-assays using internal reference. Results: The CVs of results of intra- and inter-assays on reference for precision by the prepared kit were 5.2% and 8.23% respectively. The linear range and minimum detection limit of the kit were 3.91-62.5 and 3.0 ng/ml respectively. The detection result of Y. pestis was positive, and no cross reaction with Y. pseudotuberculosis was observed. The CVs of F1 antigen at high, moderate and low contents determined by the kit were 4.54%-8.4%, while the recovery rates were 104%-108%. The kit showed high stability, of which the validity period was not less than 12 months. Conclusion: A double antibody sandwich ELISA diagnostic kit for F1 antigen of Y. pestis was successfully prepared, which provided a rapid method for clinical diagnosis and epidemiological surveillance of plaque, and a tool for determination of F1 antigen content in novel plaque component vaccine.


Chen Y.,Lanzhou Institute of Biological Products Co.
Advances in Experimental Medicine and Biology | Year: 2015

The study on dynamic analysis of human urinary proteome is the foundation that we discriminate certain various urinary proteins as potential biomarker derived from the disease itself or normal physiological change. In our results, based on RPLC-MS/MS and spectral count to study pooled and individual urine samples and other researchers’ studies, it can be known that the content of many urinary proteins maintain relatively stable. We have reason to believe that the relatively stable urinary protein is a very valuable resource as biomarkers. Many similar proteins such as prostaglandin-H2 D-isomerase and apolipoprotein D proteins have been proved our hypothesis. The following field, the number, preservation and treatment methods of urine sample, the standardization of analysis method and data processing, and suitable quantitative method, is ought to the focus of future study. © Springer Science+Business Media Dordrecht 2015.


PubMed | Lanzhou Institute of Biological Products Co.
Type: | Journal: Advances in experimental medicine and biology | Year: 2014

The study on dynamic analysis of human urinary proteome is the foundation that we discriminate certain various urinary proteins as potential bio-marker derived from the disease itself or normal physiological change. In our results, based on RPLC-MS/MS and spectral count to study pooled and individual urine samples and other researchers studies, it can be known that the content of many urinary proteins maintain relatively stable. We have reason to believe that the relatively stable urinary protein is a very valuable resource as biomarkers. Many similar proteins such as prostaglandin-H2 D-isomerase and apolipoprotein D proteins have been proved our hypothesis. The following field, the number, preservation and treatment methods of urine sample, the standardization of analysis method and data processing, and suitable quantitative method, is ought to the focus of future study.

Loading Lanzhou Institute of Biological Products Co. collaborators
Loading Lanzhou Institute of Biological Products Co. collaborators