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Antibody purification

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https://www.readbyqxmd.com/read/28101473/purification-and-characterization-of-bovine-serum-albumin-using-chromatographic-method
#1
Sanaz Balkani, Sara Shamekhi, Ramin Raoufinia, Reza Parvan, Jalal Abdolalizadeh
Purpose: Albumin is an abundant protein of blood and has many biopharmaceutical applications. The aim of this study was to purify bovine serum albumin (BSA) using produced rabbit anti-BSA antibody. Methods: The polyclonal antibody was produced against the BSA in rabbits. Then, the pure BSA was injected to three white New Zealand rabbits. ELISA test was done to evaluate antibody production. After antibody purification,the purified antibody was attached to CNBr-activated sepharose and finally it was used for purification of albumin from bovine serum...
December 2016: Advanced Pharmaceutical Bulletin
https://www.readbyqxmd.com/read/28093274/a-direct-quantitative-pcr-based-measurement-of-herpes-simplex-virus-susceptibility-to-antiviral-drugs-and-neutralizing-antibodies
#2
Dezső P Virók, Ildikó Eszik, Tímea Mosolygó, Kamil Önder, Valéria Endrész, Katalin Burián
Herpes simplex viruses (HSV) are common human pathogens that can cause painful but benign manifestations and recurrent complaints, but can also cause significant morbidity and mortality on infection of the eye or brain and with disseminated infection of an immunosuppressed patient or a neonate. HSV growth inhibition measurement by plaque or yield reduction is a key task in the development of novel antiviral compounds but the manual methods are very labour intensive. The sensitive and specific PCR technology could be an effective method for quantitation of HSV DNA related to virus replication; however the currently described PCR approaches have a major limitation, namely the requirement of purification of DNA from the infected cells...
January 13, 2017: Journal of Virological Methods
https://www.readbyqxmd.com/read/28092949/label-free-separation-of-induced-pluripotent-stem-cells-with-anti-ssea-1-antibody-immobilized-microfluidic-channel
#3
Akihisa Otaka, Kazuki Kitagawa, Takahiko Nakaoki, Mitsuhi Hirata, Kyoko Fukazawa, Kazuhiko Ishihara, Atsushi Mahara, Tetsuji Yamaoka
When induced pluripotent stem cell (iPSC) is routinely cultured, the obtained cells are a heterogeneous mixture, including feeder cells and partially differentiated cells. Therefore, a purification process is required to use them in a clinical stage. We described a label-free separation of iPSCs using a microfluidic channel. Antibodies against stage-specific embryonic antigen 1 (SSEA-1) was covalently immobilized on the channel coated with a phospholipid polymer. After injection of the heterogeneous cell suspension containing iPSCs, the velocity of cell movement under a liquid flow condition was measured...
January 16, 2017: Langmuir: the ACS Journal of Surfaces and Colloids
https://www.readbyqxmd.com/read/28092380/chromatin-immunoprecipitation-in-microfluidic-droplets-towards-fast-and-cheap-analyses
#4
Bruno Teste, Jerome Champ, Arturo Londono-Vallejo, Stéphanie Descroix, Laurent Malaquin, Jean-Louis Viovy, Irena Draskovic, Guillaume Mottet
Genetic organization is governed by the interaction of DNA with histone proteins, and differential modifications of these proteins is a fundamental mechanism of gene regulation. Histone modifications are primarily studied through chromatin immunoprecipitation (ChIP) assays, however conventional ChIP procedures are time consuming, laborious and require a large number of cells. Here we report for the first time the development of ChIP in droplets based on a microfluidic platform combining nanoliter droplets, magnetic beads (MB) and magnetic tweezers (MT)...
January 16, 2017: Lab on a Chip
https://www.readbyqxmd.com/read/28089962/mycoplasma-clearance-and-risk-analysis-in-a-model-bioprocess
#5
Julie Wang, Sarah Johnson, Matthew Brown, Scott Lute, Cyrus Agarabi, Alena Dabrazhynetskaya, Vladimir Chizhikov, Kurt Brorson
Mycoplasmas are a type of bacteria that lack cell walls and are occasional cell culture contaminants. In a biotechnology setting, because they can pass through 0.2 μm filters, mycoplasmas could pose a potential patient safety hazard if undetected contaminants from the production culture were not completely removed by downstream biotechnology manufacturing. In this study we investigated the ability of typical commercial monoclonal antibody (mAb) purification operations to clear and kill mycoplasmas, using Acholeplasma laidlawii as a model organism...
January 15, 2017: PDA Journal of Pharmaceutical Science and Technology
https://www.readbyqxmd.com/read/28089881/two-step-chromatographic-purification-of-glutathione-s-transferase-tagged-human-papillomavirus-type-16-e6-protein-and-its-application-for-serology
#6
Mei Ling Xu, Seung Cheol Kim, Hyoung Jin Kim, Woong Ju, Yun Hwan Kim, Hong-Jin Kim
Human papillomavirus (HPV) E6 protein is an oncoprotein with a pivotal role in cervical carcinogenesis. Expression and purification of HPV E6 from Escherichia coli (E. coli) has been difficult because of its strong hydrophobicity even when expressed as a fusion protein with glutathione S-transferase (GST). There has been no protocol suggested for purifying GST-tagged HPV E6 protein with high purity so far. Herein, we provide efficient protocol for purifying GST-HPV16 E6 protein for the first time. In the current study, the GST-tagged protein was expressed in E...
January 9, 2017: Protein Expression and Purification
https://www.readbyqxmd.com/read/28073086/human-neural-progenitors-derived-from-integration-free-ipscs-for-sci-therapy
#7
Ying Liu, Yiyan Zheng, Shenglan Li, Haipeng Xue, Karl Schmitt, Georgene W Hergenroeder, Jiaqian Wu, Yuanyuan Zhang, Dong H Kim, Qilin Cao
As a potentially unlimited autologous cell source, patient induced pluripotent stem cells (iPSCs) provide great capability for tissue regeneration, particularly in spinal cord injury (SCI). However, despite significant progress made in translation of iPSC-derived neural progenitor cells (NPCs) to clinical settings, a few hurdles remain. Among them, non-invasive approach to obtain source cells in a timely manner, safer integration-free delivery of reprogramming factors, and purification of NPCs before transplantation are top priorities to overcome...
January 5, 2017: Stem Cell Research
https://www.readbyqxmd.com/read/28067524/urea-artifacts-interfere-with-immuno-purification-of-lysine-acetylation
#8
Ana Martinez-Val, Fernando Garcia, Pilar Ximénez-Embún, Ailyn Martínez Teresa-Calleja, Nuria Ibarz, Isabel Ruppen, Javier Munoz
Comprehensive analysis of post-translational modifications (PTMs) often depends on the purification of modified peptides prior to LC-MS/MS. The implementation of these enrichment methods requires thorough knowledge of the experimental conditions to achieve optimal selectivity and sensitivity. In this regard, large-scale analysis of lysine acetylation, a key PTM for multiple cellular processes, makes use of monoclonal pan-antibodies designed against this moiety. We report that the immuno-purification of lysine-acetylated peptides is hampered by the copurification of lysine carbamylated peptides, a frequent urea artifact...
January 17, 2017: Journal of Proteome Research
https://www.readbyqxmd.com/read/28065273/screening-and-characterization-strategies-for-nanobodies-targeting-membrane-proteins
#9
S Veugelen, M Dewilde, B De Strooper, L Chávez-Gutiérrez
The study of membrane protein function and structure requires their successful detection, expression, solubilization, and/or reconstitution, which poses a challenging task and relies on the availability of suitable tools. Several research groups have successfully applied Nanobodies in the purification, as well as the functional and structural characterization of membrane proteins. Nanobodies are small, single-chain antibody fragments originating from camelids presenting on average a longer CDR3 which enables them to bind in cavities and clefts (such as active and allosteric sites)...
2017: Methods in Enzymology
https://www.readbyqxmd.com/read/28062629/unraveling-the-molecular-complexity-of-o-glycosylated-endogenous-n-terminal-pro-b-type-natriuretic-peptide-forms-in-blood-plasma-of-patients-with-severe-heart-failure
#10
Bernhard Halfinger, Angelika Hammerer-Lercher, Benno Amplatz, Bettina Sarg, Leopold Kremser, Herbert H Lindner
BACKGROUND: Currently, N-terminal pro-B-type natriuretic peptide (NT-proBNP) and its physiologically active counterpart, BNP, are most frequently used as biomarkers for diagnosis, prognosis, and disease monitoring of heart failure (HF). Commercial NT-proBNP and BNP immunoassays cross-react to varying degrees with unprocessed proBNP, which is also found in the circulation. ProBNP processing and immunoassay response are related to O-linked glycosylation of NT-proBNP and proBNP. There is a clear and urgent need to identify the glycosylation sites in the endogenously circulating peptides requested by the community to gain further insights into the different naturally occurring forms...
January 2017: Clinical Chemistry
https://www.readbyqxmd.com/read/28061542/an-efficient-method-for-native-protein-purification-in-the-selected-range-from-prostate-cancer-tissue-digests
#11
Rumana Ahmad, Carrie D Nicora, Anil K Shukla, Richard D Smith, Wei-Jun Qian, Alvin Y Liu
BACKGROUND: Prostate cancer (CP) cells differ from their normal counterpart in gene expression. Genes encoding secreted or extracellular proteins with increased expression in CP may serve as potential biomarkers. For their detection and quantification, assays based on monoclonal antibodies are best suited for development in the clinical setting. One approach to obtain antibodies is to use recombinant proteins as immunogen. However, the synthesis of recombinant protein for each identified candidate is time-consuming and expensive...
December 2016: Chinese Clinical Oncology
https://www.readbyqxmd.com/read/28055299/guiding-bispecific-monovalent-antibody-formation-through-proteolysis-of-igg1-single-chain
#12
Nazzareno Dimasi, Ryan Fleming, Kris F Sachsenmeier, Binyam Bezabeh, Carl Hay, Jincheng Wu, Erin Sult, Saravanan Rajan, Li Zhuang, Peter Cariuk, Andrew Buchanan, Michael A Bowen, Herren Wu, Changshou Gao
We developed an IgG1 domain-tethering approach to guide the correct assembly of two light and two heavy chains, derived from two different antibodies, to form bispecific monovalent antibodies in IgG1 format. We show here that assembling two different light and heavy chains by sequentially connecting them with protease-cleavable polypeptide linkers results in the generation of monovalent bispecific antibodies that have IgG1 sequence, structure and functional properties. This approach was used to generate a bispecific monovalent antibody targeting the epidermal growth factor receptor and the type I insulin-like growth factor receptor that: 1) can be produced and purified using standard IgG1 techniques; 2) exhibits stability and structural features comparable to IgG1; 3) binds both targets simultaneously; and 4) has potent anti-tumor activity...
January 5, 2017: MAbs
https://www.readbyqxmd.com/read/28055297/a-strategy-to-identify-linker-based-modules-for-the-allosteric-regulation-of-antibody-antigen-binding-affinities-of-different-scfvs
#13
Sarah-Jane Kellmann, Stefan Dübel, Holger Thie
Antibody single-chain variable fragments (scFvs) are used in a variety of applications, such as for research, diagnosis and therapy. Essential for these applications is the extraordinary specificity, selectivity and affinity of antibody paratopes, which can also be employed for efficient protein purification. However, this use is hampered by the high affinity for the protein to be purified because harsh elution conditions, which may impair folding, integrity or viability of the eluted biomaterials, are typically required...
January 5, 2017: MAbs
https://www.readbyqxmd.com/read/28043587/cloning-expression-and-purification-of-recombinant-protein-mpt-64-from-a-virulent-strain-of-mycobacterium-bovis-in-a-prokaryotic-system
#14
Maryam Mohammadi Tashakkori, Majid Tebianian, Mohammad Tabatabaei, Nader Mosavari
OBJECTIVE: Tuberculosis (TB) is a zoonotic infectious disease common to humans and animals that is caused by the rod-shaped acid-fast bacterium Mycobacterium bovis. Rapid and sensitive detection of TB is promoted by specific antigens. Virulent strains of the TB complex from M. bovis contain 16 regions of difference (RD) in their genome that encode important proteins, including major protein of Mycobacterium Tuberculosis 64 (MBT-64, which is a primary immune-stimulating antigen encoded by RD-2...
December 2016: International Journal of Mycobacteriology
https://www.readbyqxmd.com/read/28043570/evaluate-the-efficiency-of-antigen-60-a60-protein-from-bcg-strain-of-mycobacterium-bovis-as-a-diagnostic-antigen
#15
Nafiseh Shakibamehr, Nader Mosavari, Mahdi Babaie, Samerand Reshady
OBJECTIVE/BACKGROUND: Tuberculosis (TB) is one of the most common infectious diseases in Iran and around the world. Diagnosis of this disease in many cases is difficult and often requires the use of paraclinical methods. Current diagnostic methods are either too slow or lack enough sensitivity or specificity. Several mycobacterial antigens are involved in the complex interaction with the immune system of the host. They can be helpful for mycobacteria diagnosis. Antigen 60 (A60) is a thermostable antigen found in the cytosol of Mycobacterium bovis and Mycobacterium tuberculosis...
December 2016: International Journal of Mycobacteriology
https://www.readbyqxmd.com/read/28042093/expression-and-antigenicity-of-recombinant-human-respiratory-syncytial-virus-glycoproteins-having-different-affinity-tags
#16
Han Saem Lee, A-Reum Kim, Kisoon Kim, Wan-Ji Lee, Sung Soon Kim, You-Jin Kim
Human respiratory syncytial virus (HRSV) is a main cause of lower respiratory tract infections in infants and the elderly. Glycoprotein (G) is major antigen on the viral surface, and plays a key role for virus entry. Therefore, purification of the glycoprotein of HRSV is critical for the development of HRSV vaccine and serological diagnosis. In this study, we report the design and characterization of glycoprotein engineered rationally to enhance the protein solubility and to facilitate efficient purification...
December 29, 2016: Protein Expression and Purification
https://www.readbyqxmd.com/read/28025125/production-purification-and-immunogenicity-of-recombinant-ebola-virus-proteins-a-comparison-of-freund-s-adjuvant-and-adjuvant-system-03
#17
Krister Melén, Laura Kakkola, Felix He, Kari Airenne, Olli Vapalahti, Helen Karlberg, Ali Mirazimi, Ilkka Julkunen
There is an urgent need for Ebola virus (EBOV) proteins, EBOV-specific antibodies and recombinant antigens to be used in diagnostics and as potential vaccine candidates. Our objective was to produce and purify recombinant proteins for immunological assays and for the production of polyclonal EBOV specific antibodies. In addition, a limited comparison of the adjuvant effects of Freund's complete adjuvant (FCA) and adjuvant system 03 (AS03) was carried out. Recombinant EBOV GST-VP24, -VP30, -VP35, -VP40 and -NP were produced in E...
December 24, 2016: Journal of Virological Methods
https://www.readbyqxmd.com/read/28012937/affinity-chromatography-a-versatile-technique-for-antibody-purification
#18
REVIEW
Sushrut Arora, Vikas Saxena, B Vijayalakshmi Ayyar
Antibodies continue to be extremely utilized entities in myriad applications including basic research, imaging, targeted delivery, chromatography, diagnostics, and therapeutics. At production stage, antibodies are generally present in complex matrices and most of their intended applications necessitate purification. Antibody purification has always been a major bottleneck in downstream processing of antibodies, due to the need of high quality products and associated high costs. Over the years, extensive research has focused on finding better purification methodologies to overcome this holdup...
December 22, 2016: Methods: a Companion to Methods in Enzymology
https://www.readbyqxmd.com/read/28012750/purification-of-a-human-prostate-specific-antigen
#19
M C Wang, L A Valenzuela, G P Murphy, T M Chu
Rabbit antiserum raised against the crude extract of normal human prostatic tissue contained antibodies to a prostatic tissue-specific antigen as shown by immunoprecipitation techniques. Using this antiserum a prostate antigen was detected in normal, benign hypertrophic, and malignant prostatic tissues, but not in other human tissues. The prostate antigen was purified to homogeneity from prostatic tissues and showed a single protein band on analytical polyacrylamide gel electrophoresis and isoelectric focusing...
December 21, 2016: Journal of Urology
https://www.readbyqxmd.com/read/28010734/proteins-adopt-functionally-active-conformations-after-type-iii-secretion
#20
Kevin James Metcalf, James Lea Bevington, Sandy Lisette Rosales, Lisa Ann Burdette, Elias Valdivia, Danielle Tullman-Ercek
BACKGROUND: Bacterial production of natively folded heterologous proteins by secretion to the extracellular space can improve protein production by simplifying purification and enabling continuous processing. In a typical bacterial protein production process, the protein of interest accumulates in the cytoplasm of the cell, requiring cellular lysis and extensive purification to separate the desired protein from other cellular constituents. The type III secretion system of Gram-negative bacteria is used to secrete proteins from the cytosol to the extracellular space in one step, but proteins must unfold during translocation, necessitating the folding of secreted proteins in the extracellular space for an efficient production process...
December 23, 2016: Microbial Cell Factories
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