99 results about "Mammalian expression" patented technology

The present invention relates to the recombinant method used for the production of soluble form of an antibody that binds to CD20 for treatment of patients with relapsed or refractory, low-grade or follicular, CD20-positive, B-cell non-Hodgkin's lymphoma (NHL). The treatment will comprise the use of immunologically active anti-CD20 antibodies; or radiolabeled anti-CD20 antibodies and or cooperative strategies where both labeled and non-labeled antibodies will be used for treatment of NHL. The procedure describes the de novo synthesis of the nucleic acid sequence encoding anti-CD20, transformation of the constructed nucleic acid sequences into competent bacteria and the sub-cloning of the same into mammalian expression vectors for expression of the desired protein. DNA constructs comprising the control elements associated with the gene of interest has been disclosed. The nucleic acid sequence of interest has been codon optimized to permit expression in the suitable mammalian host cells.

The present invention provides a process and methods for producing asymmetric antibodies in a mammalian expression system. The asymmetric antibodies are transiently or stably expressed and in cells that stably express the asymmetric antibody, following a rapid 2-step process of stable pool to clone, a highly pure asymmetric antibody expressing clone can be identified at a success frequency that permits for screening of tens of clones rather than thousands. The asymmetric antibodies are produced at a high titre and with a high level of purity with no contaminating homodimer antibodies following protein A purification with a step yield of near 100%. Typical downstream purification processes employ standard hydrophobic interaction chromatography (HIC) and/or cation exchange (CEX) resins and the antibody is stable within a wide dynamic range of buffer pH (4-8) and within the requirements for manufacturing antibodies for pre-clinical and clinical applications.

A method for measuring HCV NS3/4A activity from a HCV NS3/4A sequence, comprising obtaining and cloning the sequence into a mammalian expression vector, transiently transfecting a mammalian cell with the vector, which includes a reporter construct encoding a HCV NS3/4A cleavage site fused to a detectable label, measuring signal production from the label resulting from cleavage at the cleavage site, and measuring effects on signal production by addition of a test compound.

A construct comprising the ectodomain of the Hepatitis C Virus (HCV) E2 sequence and a mammalian expression system therefor is disclosed. The construct comprises a CMV promoter, prolactin signal sequence, the ectodomain of HCV E2 sequence truncated at aa 664, a thrombin cleavage site and a human Fc domain. The method also relates to an expression system for the construct, which is stably expressed in human embryonic kidney cells 293T. Continuous protein expression in a bioreactor allows for 4 mg of purified protein per liter of cell supernatant.

The invention discloses an efficient clone selection expression vector, which is obtained by an ITRs sequence of a piggyBac transposition system inserted into a mammal expression vector. The invention also discloses a preparation method and use thereof. The efficient clone selection expression vector is built by using the ITRs sequence of the piggyBac transposition system, and meanwhile, a transcriptase gene which is optimized by a codon and can be efficiently expressed in mice is cloned to another eukaryotic expression vector. A selection marker and a target gene are integrated into a genome of a host cell in common by coexpression of an expression vector containing the ITRs sequence and the vector containing transcriptase. Thus, the transfection efficiency and the positive clone screening rate are greatly improved.

An improved mammalian expression vector system which allows: (1) highly expressing exogenous proteins in host mammalian cells; (2) rapidly and efficiently screening recombinant stable cell lines expressing the gene of interest; (3) maintain sustainable expression and prevent gene silencing; and (4) effectively secreting proteins into media in some of cases. The entire expression vector system includes optimized promoters and core promoters, the use of special internal ribosome entry sites, integration of the bacterial backbone into the mammalian expression unit, multiple choices of selection markers, artificial matrix attachment region elements, effective secreting lead sequences and their 5′ and 3′ UTRs, and proper combinations of these expression elements.

The present invention relates to the use of internal promoters in mammalian expression vectors including plasmid vectors and enhancer-deleted retroviral vectors. The retroviral vectors have improved safety and optimal levels of transgene expression and vector titers.

The present invention relates to protein expression systems and in particular to a mammalian protein expression system. Specifically, the present invention provides a rodent cell line with enhanced protein production capabilities. The invention also relates to eukaryotic cloning and expression vectors and related methods, and in particular to DNA vectors capable of high level expression of a protein of interest. The invention allows for long-term episomal maintenance of expression vectors in mammalian cells.

The present invention relates to the recombinant method used for the production of soluble form monoclonal antibody that binds to CD52. The procedure describes the de novo synthesis of the nucleic acid sequence encoding anti-CD 52, transformation of the constructed nucleic acid sequences into competent bacteria and the sub-cloning of the same into mammalian expression vectors for expression of the desired protein. DNA constructs comprising the control elements associated with the gene of interest has been disclosed. The nucleic acid sequence of interest has been codon optimized to permit expression in the suitable mammalian host cells.

The invention provides methods for generating high titers of high-affinity antibodies from hybridoma cells produced by fusing myeloma cells with in vitro immunized donor cells. The hybridoma cells or mammalian expression cells with cloned antibody genes from the hybridomas producing the high-affinity antibodies may be mismatch repair defective due to defects of endogenous mismatch repair subunits of through expression of a dominant negative allele of a mismatch repair gene which allows the hybridoma cell to be hypermutable, may be rendered hypermutable by chemical means, or may be naturally mismatch repair deficient. High-affinity antibodies and high titer producer cells producing antibodies may be prepared by the methods of the invention.

The present invention features mammalian expression systems with improved production yields, and method of using these systems to produce desired proteins. In one embodiment, the expression systems of the present invention comprise genetically-engineered mammalian host cells cultured in a medium that contains an effective amount of heparin or heparin-like molecules. The presence of heparin or heparin-like molecules significantly increases protein production by the cultured cells. The present invention also features the use of constitutively-active components of FGFR-I-mediated signal transduction pathways to improve protein production by cultured mammalian cells. Co-expression of such a component with a protein of interest markedly increases the production yield of the protein of interest.

The invention discloses a gene expression regulation and control method and system based on Type I-F CRISPR/ Cas. The first study shows that a Cascade compound, such as a PaeCascade compound, belonging to a Type I-F CRISPR/ Cas system can be efficiently combined with a target site in a mammalian cell; and meanwhile, the Type I-F CRISPR/ Cas system is subjected to mammalian expression system optimization, so that a transcription activation factor or a transcription inhibition factor can be efficiently recruited to the target site in the mammalian cell, and a transcription activation system anda transcription inhibition system are successfully constructed. The application of the Type I-F CRISPR/ Cas system in the aspect of gene regulation and control of the mammalian cell becomes possible,and a necessary tool is provided for gene transcription regulation and control based on the Type I-F CRISPR/ Cas system.

The invention provides a preparation method of dendritic cells, which belongs to the technical field of tumor treatment. The preparation method comprises the steps of: 1) ligating a fusion gene of human leukocyte antigen (HLA) and KRAS mutation to pCDH or a mammalian expression vector to obtain a recombinant plasmid; 2) transforming the dendritic cells (DC) derived from peripheral blood by electroporation with the recombinant plasmid to obtain the recombinant cells (i-DC); and 3) inducing maturation of the recombinant cells to obtain the mature dendritic cells. In the prior art, antigen-presenting cells are generally loaded by mutating polypeptides, or mutant antigens are transferred into antigen-presenting cells, but the efficiency of both is low, The method converts the fusion gene of KRAS antigen and HLA into the antigen-presenting cells and overexpresses two genes, increases the binding rate of HLA and KRAS mutant antigen, improves the efficiency of presentation, and more easily obtains CTL with specific killing effect.