全文获取类型
收费全文 | 89篇 |
免费 | 0篇 |
专业分类
89篇 |
出版年
2013年 | 2篇 |
2011年 | 2篇 |
2009年 | 1篇 |
2008年 | 3篇 |
2007年 | 4篇 |
2006年 | 9篇 |
2005年 | 6篇 |
2004年 | 6篇 |
2003年 | 6篇 |
2002年 | 2篇 |
2001年 | 3篇 |
2000年 | 8篇 |
1999年 | 3篇 |
1998年 | 9篇 |
1997年 | 1篇 |
1996年 | 1篇 |
1993年 | 1篇 |
1992年 | 1篇 |
1991年 | 1篇 |
1990年 | 2篇 |
1989年 | 2篇 |
1988年 | 1篇 |
1987年 | 1篇 |
1980年 | 1篇 |
1979年 | 1篇 |
1978年 | 1篇 |
1977年 | 1篇 |
1974年 | 1篇 |
1972年 | 1篇 |
1971年 | 1篇 |
1969年 | 3篇 |
1968年 | 1篇 |
1967年 | 2篇 |
1966年 | 1篇 |
排序方式: 共有89条查询结果,搜索用时 15 毫秒
1.
Production of Fab fragments of monoclonal antibodies using polyelectrolyte complexes 总被引:4,自引:0,他引:4
Dainiak MB Muronetz VI Izumrudov VA Galaev IY Mattiasson B 《Analytical biochemistry》2000,277(1):58-66
A new method for the production of monovalent Fab fragments of antibodies has been developed. Traditionally Fab fragments are produced by proteolytic digestion of antibodies in solution followed by isolation of Fab fragments. In the case of monoclonal antibodies against inactivated subunits of glyceraldehyde-3-phosphate dehydrogenase, digestion with papain resulted in significant damage of the binding sites of the Fab fragments. Antigen was covalently attached to the polycation, poly(N-ethyl-4-vinylpyridinium bromide). Proteolysis of monoclonal antibodies in the presence of the antigen-polycation conjugate followed by (i) precipitation induced by addition of polyanion, poly(methacrylic) acid, and pH shift from 7.3 to 6.5 and (ii) elution at pH 3.0 resulted in 90% immunologically competent Fab fragments. Moreover, the papain concentration required for proteolysis was 10 times less in the case of antibodies bound to the antigen-polycation conjugate than that of free antibodies in solution. The digestion of antibodies bound to the antigen-polyelectrolyte complex was less damaging, suggesting that binding to the antigen-polycation conjugate not only protected binding sites of monoclonal antibodies from proteolytic damage but also facilitated the proteolysis probably by exposing antibody molecules in a way convenient for proteolytic attack by papain. 相似文献
2.
Noppe W Plieva FM Vanhoorelbeke K Deckmyn H Tuncel M Tuncel A Galaev IY Mattiasson B 《Journal of biotechnology》2007,131(3):293-299
Selected phage clones expressing a peptide with high binding affinity for recombinant human lactoferrin or von Willebrand factor (vWF) were covalently coupled to macroporous poly(dimethylacrylamide) monolithic column. Large pore size (10-100 microm) of macroporous poly(dimethylacrylamide) makes it possible to couple long (1 microm) phage particles as ligands without any risk of blocking the monolithic column. The macroporous monolithic columns were successfully used for the direct affinity capture of target proteins from particulate containing feeds like milk containing casein micelles and fat globules (1-10 microm in size) or even whole blood containing blood cells (up to 20 microm in size). The newly developed platform based on selected bacteriophages immobilized within macropores of the monolithic cryogels presents a convenient alternative to antibodies for fast and selective development of the specific adsorbent. 相似文献
3.
Lac repressor protein was purified from E. coli BMH8117 harboring plasmid pWB1000 and E. coli K12BMH 71-18 strains. Displacement of the protein with poly(ethyleneimine) (PEI) from phosphocellulose cation exchange column was shown to be an effective elution strategy. It resulted in better recoveries and sharper elution profiles than traditional salt elution without effecting the purity of the protein. The elution is assumed to proceed via displacement of bound protein by PEI when the polymer binds to the ion exchanger. The minor impurities in the protein solution were finally removed by chromatography on immobilized metal affinity column. The repressor protein undergoes distinct conformational changes upon addition of specific inducer isopropyl--D-thiogalactoside (IPTG), which is evidenced by changes in ultraviolet absorption spectrum. The protein was immobilized covalently to the Sepharose matrix. The intact biological activity of the protein after immobilization was shown by binding of genomic DNA and lac operator plasmid DNA from E. coli to the immobilized lac repressor. 相似文献
4.
Wahlund PO Gustavsson PE Izumrudov VA Larsson PO Galaev IY 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》2004,807(1):121-127
The demand for highly purified plasmids in gene therapy and plasmid-based vaccines requires large-scale production of pharmaceutical-grade plasmid. Plasmid DNA was selectively precipitated from a clarified alkaline lysate using the polycation poly(N,N'-dimethyldiallylammonium) chloride which formed insoluble polyelectrolyte complex (PEC) with the plasmid DNA. Soluble PECs of DNA with polycations have earlier been used for cell transformation, but now the focus has been on insoluble PECs. Both DNA and RNA form stable PECs with synthetic polycations. However, it was possible to find a range of salt concentration where plasmid DNA was quantitatively precipitated whereas RNA remained in solution. The precipitated plasmid DNA was resolubilised at high salt concentration and the polycation was removed by gel-filtration. 相似文献
5.
Balan S Murphy J Galaev I Kumar A Fox GE Mattiasson B Willson RC 《Biotechnology letters》2003,25(13):1111-1116
The affinity of metal chelates for amino acids, such as histidine, is widely used in purifying proteins, most notably through six-histidine `tails'. We have found that metal affinity interactions can also be applied to separation of single-stranded nucleic acids through interactions involving exposed purines. Here we describe a metal affinity precipitation method to resolve RNA from linear and plasmid DNA. A copper-charged copolymer of N-isopropyl acrylamide (NIPAM) and vinyl imidazole (VI) is used to purify plasmid from an alkaline lysate of E. coli. The NIPAM units confer reversible solubility on the copolymer while the imidazole chelates metal ions in a manner accessible to interaction with soluble ligands. RNA was separated from the plasmid by precipitation along with the polymer in the presence of 800 mM NaCl. Bound RNA could be recovered by elution with imidazole and separated from copolymer by a second precipitation step. RNA binding showed a strong dependence on temperature and on the type of buffer used. 相似文献
6.
Ivanov AE Edink E Kumar A Galaev IY Arendsen AF Bruggink A Mattiasson B 《Biotechnology progress》2003,19(4):1167-1175
Conjugation of penicillin acylase (PA) to poly-N-isopropylacrylamide (polyNIPAM) was studied as a way to prepare a thermosensitive biocatalyst for industrial applications to antibiotic synthesis. Condensation of PA with the copolymer of NIPAM containing active ester groups resulted in higher coupling yields of the enzyme (37%) compared to its chemical modification and copolymerization with the monomer (9% coupling yield) at the same NIPAM:enzyme weight ratio of ca. 35. A 10-fold increase of the enzyme loading on the copolymer resulted in 24% coupling yield and increased by 4-fold the specific PA activity of the conjugate. Two molecular forms of the conjugate were found by gel filtration on Sepharose CL 4B: the lower molecular weight fraction of ca. 10(6) and, presumably, cross-linked protein-polymer aggregates of MW > 10(7). Michaelis constant for 5-nitro-3-phenylacetamidobenzoic acid hydrolysis by the PA conjugate (20 microM) was found to be slightly higher than that of the free enzyme (12 microM), and evaluation of V(max) testifies to the high catalytic efficiency of the conjugated enzyme. PolyNIPAM-cross-linked PA retained its capacity to synthesize cephalexin from d-phenylglycin amide and 7-aminodeacetoxycephalosporanic acid. The synthesis-hydrolysis ratios of free and polyNIPAM-cross-linked enzyme in cephalexin synthesis were 7.46 and 7.49, respectively. Thus, diffusional limitation, which is a problem in the industrial production of beta-lactam antibiotics, can be successfully eliminated by cross-linking penicillin acylase to a smart polymer (i.e., polyNIPAM). 相似文献
7.
In the previous work, after screening tropical plants (43 species) for peroxidase activity, high activity has been detected in leaves of some palms and especially African oil palm Elaeis guineensis. This palm is widely cultivated in Colombia and presents a promising source for the industrial production of peroxidase. The initial enzyme isolation included homogenization and extraction of pigments using aqueous two phase polymer system. Initially, traditional system, formed by polyethyleneglycol/K2HPO4, was used. The replacement of K2HPO4 with (NH4)2SO4 allowed direct application of the salt phase with accumulated peroxidase on a Phenyl-Sepharose column. The final purification was carried out by liquid chromatography on Sephacryl S200 and DEAE-Toyopearl columns. The specific activity of the purified peroxidase measured toward guaiacol was 4300 units per mg of protein. The molecular weight and isoelectric point for palm peroxidase were 57.000 and 3.8, respectively. Palm peroxidase possesses uniquely high thermostability and is more stable in organic solvents than horseradish peroxidase is. 相似文献
8.
Glycoproteins, as a class of biomolecules, exhibit much more heterogeneous structures than non-glycosylated proteins. They present a challenging area of research. Model glycoproteins with well-defined protein and carbohydrate structures are helpful in the search for high-resolution methods for the separation of glycoproteins. Neoglycoproteins, maltose-modified chymotrypsin and lactose-modified chymotrypsin, were synthesised by modifying chymotrypsin with maltose and lactose, respectively, using the reductive amination method. Boronate chromatography was applied to isolate the neoglycoproteins from non-glycosylated substances. The use of Tris–HCl as a shielding reagent during the boronate chromatography proved to be efficient in eliminating unwanted interactions between the boronate ligand and the peptide backbone of chymotrypsin. The retention time of neoglycoproteins on the boronate column was increased with increasing the degree of modification. 相似文献
9.
Wheat-aegilops hybrid plants Triticum aestivum L. (2n = 42) x Aegilops cylindrica Host (2n = 28) were investigated with using microsatellite markers. In two BC1F9 lines some genome modifications connected with losing DNA fragments of initial variety or appearing of Aegilops genome elements were detected. In some investigated hybrids new amplicons lacking in parental plants were found. Substitution of wheat chromosomes for aegilops chromosomes was not revealed. Analysis of microsatellite loci in BC2F5 plants showed stable introgression of aegilops genetic elements into wheat; elimination of some transferred aegilops DNA fragments in the course of backcrossing; decreasing size of introgressive elements after backcrossing. Introgressive lines were classified according to genome changes. 相似文献
10.
Kumar A Rodríguez-Caballero A Plieva FM Galaev IY Nandakumar KS Kamihira M Holmdahl R Orfao A Mattiasson B 《Journal of molecular recognition : JMR》2005,18(1):84-93
The capture of human acute myeloid leukemia KG-1 cells expressing the CD34 surface antigen and the fractionation of human blood lymphocytes were evaluated on polyvinyl alcohol (PVA)-cryogel beads and dimethyl acrylamide (DMAAm) monolithic cryogel with immobilized protein A. The affinity ligand (protein A) was chemically coupled to the reactive PVA-cryogel beads and epoxy-derivatized monolithic cryogels through different immobilization techniques and the binding efficiency of the cell surface receptors specific antibody-labeled cells to the gels/beads was determined. The binding of cells to monolithic cryogel was higher (90-95%) compared with cryogel beads (76%). B-lymphocytes, which bound to the protein A-cryogel beads, were separated from T-lymphocytes with yields for the two cell types 74 and 85%, respectively. About 91% of the bound B-cells could be recovered without significantly impairing their viability. Our results show differences in the percentage of cell-binding to the immunosorbents caused by ligand density, flow shear forces and bond strength between the cells and the affinity surface once distinct chemical coupling of protein A, size of beads, sequence of antibody binding to protein A adsorbents, morphology and geometry of surface matrices were compared. 相似文献