Creative Biolabs is one of the well-recognized experts who is professional in applying advanced phage display technologies for a broad range of project objectives. With years of experience, our scientists can offer high-quality phage display library construction and custom phage display library screening services to meet our clients’ demands precisely. Particularly, our services also involve specific antibody discovery (e.g. PTM-specific antibody, anti-idiotype antibody, and agonistic antibody) and peptidome discovery.

Library Construction and Screening

Fig.1 Workflow of phage display construction
and screening (Díez et al. 2015)

Phage display is one of the most powerful and widely used laboratory technique for the study of protein-protein, protein-peptide and protein-DNA interactions. This technology is mainly based on displaying the interest protein (peptides, antibodies, scaffolds or others) on the surface of employing phage and then be used to interrogate the constructed libraries containing millions or even billions of displayed phages. Theoretically, phage display is an exogenous gene expression method which the gene encoding the interest protein is inserted into bacteriophage coat protein gene then displaying the interest protein on the phage surfaces, resulting in a connection between genotype and phenotype.

Due to the different properties and advantages, Creative Biolabs is pleased to tailor the most appropriate phage display system (M13, T4 or T7) to meet our customers’ demands. Through DNA manipulation, numerous gene variants can be created and constructed as phage display library. Our scientists are confident in generating high-quality libraries with the diversity of 10⁸. Furthermore, to detect the interaction between displayed protein and those other molecules or isolate specific target binders, Creative Biolabs also utilizes the novel biopanning strategies to select high-affinity binders or ligands which are able to recognize the naïve targets.

Features

• Immune, naïve or synthetic library construction
• Pre-made library screening
• Large library capacity from 10⁷ to over 10¹²
• High affinity from 10^-7 to 10^-9
• Various phage display systems (M13, T4, T7)
• Tailored biopanning strategies
• Comprehensive QC validation

Antibody Phage Display Library

Antibody libraries are constructed by the genomic information coding for antibody variable domains, which can be derived from B cells of immune or naïve donors. Antibodies are the first proteins which were successfully displayed on the surface of phage by fusing the coding sequence of scFv or Fab to the coat protein. Due to the smaller size, scFv libraries are genetically more stable than Fab libraries, while Fab libraries lack the tendency to form higher molecular weight species, such as dimers and trimers, which can simplify the selection and characterization. Through binding the specific target, the interest antibody can then be eluted and retrieved.

Compared with the traditional hybridoma method, antibody phage display library has distinct advantages on discovering novel monoclonal antibodies and even the fully human antibody. Creative Biolabs is able to construct immune antibody libraries and isolate monoclonal antibodies with high specificity and affinity from a comprehensive list of species, which including but not limited to monkey, llama, camel, shark, alligator, mouse, rat, hamster, guinea pig, rabbit, chicken, dog, bovine, goat, sheep, and ferret. In addition, our scientists also possess some high-quality pre-made libraries (including human antibody libraries) for our library screening services.

Creative Biolabs has long-term devoted to the development and application of phage display technology. With years of experience, our scientists have developed several phage display based platforms and tailored hundreds of particular libraries and thousands of specific antibody products to boost our global customers’ research and project goals. We are pleased to use our extensive experience and advanced platform to offer the best service and the most qualified products to satisfy each demand from our customers.

Reference

1. Díez, P., Jara-Acevedo, R., González-González, M., Casado-Vela, J., Dasilva, N., Lécrevisse, Q., Bartolomé, R., Claros, J. C., González, A. and López, R. (2015) ‘High-throughgput phage-display screening in array format’, Enzyme and microbial technology, 79, 34-41.

In terms of the extensive experience in antibody research, Creative Biolabs is proud to offer our clients a series of single domain antibody services with the best quality and most competitive price.

What is Single Domain Antibody?

Single domain antibody, also known as domain antibody, V_HH, V_NAR or sdAb, is a kind of antibody fragments consisting of a single monomeric variable antibody domain and lacking the light chain and CH domain of the heavy chain in conventional Fab region. The first single domain antibody was engineered from the V_HH domain of heavy-chain antibody identified in camelids (e.g. dromedaries, camels, llamas, and alpacas). And the V_NAR domain of cartilaginous fishes (e.g. shark) heavy-chain antibody (known as IgNAR, immunoglobulin new antigen receptor) is another qualified source for sdAb development. An alternative approach is to split the dimeric variable domains from normal IgG of humans or mice into monomers by camelizing a few critical residues.

Fig.1 Distinguishing structural features of camelid and shark hcAbs. (Wesolowski et al. 2009)

Similar to a conventional antibody, single domain antibody is able to bind selectively to a specific antigen. In terms of only 12-15 kDa molecular weight, which is much smaller than either full length antibody (150-160 kDa) or other antibody fragments (Fab ~50 kDa, scFv ~25 kDa), single domain antibody takes great advantages of stability and penetrability, which are essential to the development of several antibody drugs or diagnostic tools.

Advantages of Single Domain Antibody

• High stability to function and exist within extreme conditions
• Outstanding penetrability which is able to cross the blood-brain barrier
• Recognize novel epitopes that regular size antibodies cannot
• Improve bioavailability for pharmaceutical applications
• Great potential in downstream engineering (bispecific antibody, humanization)
• Can be expressed in both eukaryotic and prokaryotic systems

Creative Biolabs is a well-recognized leader in the field of single domain antibody. Based on our advanced phage display technology, our scientists are specialized in the production and discovery of specific single domain antibodies. According to our client’s demands, we can immunize camel, llama, alpaca or even shark as repertoire source to develop single domain antibodies against interested targets (antigens). Our advanced technology allows various targets to be employed in immunization, including large protein molecules, membrane proteins, haptens and other challenging targets. As an alternative, Creative Biolabs can also screen our pre-made HuSdL® Human Single Domain Antibody Library to isolate high-potency camelized human single domain antibodies. In addition to our sdAb related services, Creative Biolabs also provides a comprehensive list of recombinant single domain antibody products to meet our customers’ requirements.

Featured sdAb Related Services

• Single Domain Antibody Library Construction and Screening
• Intrabody Discovery
• Premade Single Domain Antibody Library Screening
• Bispecific Single Domain Antibody Production
• Anti-membrane protein sdAb Production
• Anti-idiotypic sdAb Production
• Anti-BBB sdAb Production
• Anti-Albumin Single Domain Antibody Discovery Service
• Single Domain Antibody Humanization
• Single Domain Antibody Conjugation Services
• sdAb Affinity Maturation
• Single Domain Antibody Stability Improvement Service
• De Novo sdAb Sequencing
• Antibody Camelization

According to the variety of unique features, single domain antibodies can be used as potential tools for academic research, diseases diagnosis and treatment, and biotechnology development. Creative Biolabs is pleased to share our cutting-edge technology and extensive expertise in single domain antibody to facilitate our clients’ research and project development. We can offer high-quality customized services by adjusting protocols to meet even the most specific requirements.

Reference

1. Wesolowski, J., Alzogaray, V., Reyelt, J., Unger, M., Juarez, K., Urrutia, M., Cauerhff, A., Danquah, W., Rissiek, B. and Scheuplein, F. (2009) ‘Single domain antibodies: promising experimental and therapeutic tools in infection and immunity’, Medical microbiology and immunology, 198(3), 157-174.

Hybridoma technology is a technology of forming hybrid cell lines (hybridomas) by fusing a specific antibody-producing B cell with a myeloma (B cell cancer) cell that is selected for its ability to grow in tissue culture and for an absence of antibody chain synthesis. The antibodies produced by the hybridoma are all of a single specificity and are therefore monoclonal antibodies.

Creative Biolabs provides a full range of services in custom polyclonal and monoclonal antibody production from gene expression or peptide synthesis to antibody purification and labeling. We have the capacity of producing antibodies in either research quantities or in large scale for diagnostic industry. We do epitope prediction, peptide synthesize, protein expression, polyclonal or monoclonal antibody production as well as antibody purification from cell culture, ascites fluid or whole serum. We also prepare mono-specific polyclonal antibodies by ligand affinity chromatography. Importantly, we can produce monoclonal antibodies within 70 days. Creative BioLabs also has the expertise in handling antigen insufficiency and targeting very impure antigens.

Creative Biolabs has established standard operating procedures in selectively identifying antibodies that bind not only to immunogens but also to the corresponding target antigens in ELISA, Western blotting, Immuno-precipitation, Immuno-fluorescence and Immunohistochemistry. Frequently, polyclonal or monoclonal antibodies raised using peptides and recombinant proteins cannot recognize the target antigens (e.g. a protein antigen in tissue) and are essentially useless. To generate antibodies that recognize a native antigen, we employ or predict the 3-D structure of the target protein and design peptide immunogens that display on the protein surface and contain linear continuous epitopes. If a recombinant protein immunogen is used, we usually express it in a soluble form to retrieve the immunogenic properties of its native counterpart. Finally, we use our well-tested ELISA, Western blotting, IP, IF and IHC screening protocols to validate (monoclonal) antibodies that recognize the natural target antigen.

Features of the Services:
• Specialty in generating IHC-specific antibodies
• Monoclonal antibody production in 70 Days
• Expertise in antigen impurity and insufficiency
• Industry-scale antibody production
• The industry’s lowest price

As an industrial leader specialized in custom services, Creative Biolabs also provides gene-engineered antibodies including single chain and humanized antibodies. Moreover, we can conduct phage display antibody library screening and provide monoclonal Fab/ScFv antibodies derived from phage display libraries. We also provide custom IgY production services.

Creative Biolabs has proudly developed the unique Native™ Antibody Discovery Platform to develop native monoclonal antibodies using antigen-specific B lymphocytes cytometry technology.

Current in vitro antibody display methods cannot provide antibodies with naturally paired heavy chain and light chain. For technical reasons, the natural pairings of heavy and light chains found in antibody-producing B cells are not retained as the combinatorial libraries are constructed, leading to a random pairing of VH and VL in those de novo or naïve human antibody libraries. This could potentially lead to the generation of antibody molecules that have suboptimal solubility or other biophysical characteristics. Thereby, antibodies derived from these technologies most likely do not represent the naturally cognate heavy and light chains.

Another limitation often observed is that antibody derived from in vitro antibody display methods generally have moderate affinity to the targets, often in the 1-50 nM KD range, so they often require further affinity maturation before used as potential therapeutics. This is due to a practical limitation to the library size and hence diversity that can be constructed. Phage libraries are restricted to about 10¹⁰ to 10¹¹ total diversity by the efficiency of bacterial transformation as well as by the practical limit of phage particle density of about 10¹³/mL.

To solve this problem, Creative Biolabs has developed the exclusive Native™ Antibody Discovery Platform to isolate single antigen-specific plasma cells to retrieve native antibodies with extremely high in vivo specificity and affinity. Moreover, this new technology allows us to select a large number of antibody candidates at once. In addition, the selection only takes 2-3 weeks post-immunization. It should be considered by far the most efficient way in developing native monoclonal antibodies from humans, rabbits, rats, mice, chicken and other species such as camelids, sheep, monkeys, dogs and even cows. For initiating native antibody development, single B cells need to be isolated and identified. Based on the principles used, the existing techniques can be classified into two groups, the one based on cell physical properties and the other one based on cellular biological characteristics.

Main methods for single cell isolation in native antibody development:

• FACS (fluorescent activated cell sorting)
• LCM (laser-capture microdissection)
• Microengraving
• Droplet microfluidics

Fig 1. Some single cell isolation technologies (Ping Hu, et al. 2016)

Advantages of our Native™ Antibody Discovery Service including but not limited to:

• Native paired heavy chain and light chain
• In vivo affinity maturation
• High specificity and stability in vivo
• Large pool of antibody candidates
• Competitive and affordable price
• Diverse species available

Diverse species that can be used to develop native antibody

As a pioneer and the undisputed global leader in antibody discovery and manufacturing, Creative Biolabs offers native monoclonal antibodies for both academic and clinical use. Our highly professional scientists are confident in providing the domestic and international customers with the best possible services while at the most competitive price.

As a pioneer and the undisrupted global leader in antibody discovery and engineering, Creative Biolabs proudly introduces our professional service of anti-membrane protein antibody discovery with our diverse technologies.

Membrane proteins, which consist of 20~30% of total encoding genes in the genome, play a critical role in multiple biological processes, including ligand-receptor binding, signal transduction, molecule transportation, intercellular recognition, and cell adhesion. Currently, there rises growing interest regarding membrane proteins as promising targets for various drug development and other applications.

To obtain highly functional anti-membrane protein antibodies, expression and presentation of the membrane protein of interest as immunogen is the prerequisite. As most membrane proteins are partially or completely embedded in the lipid bilayer, traditional expression methods meet obstacles in presenting the native conformations, such as low expression level, difficult extraction, misfolding, and incomplete post-translational modifications. With a full understanding of the critical requirements in this field, Creative Biolabs has developed a novel Magic™ platform, which enables high-efficient expression of membrane proteins and following antibody development routes based on our multiple impeccable technologies.

For membrane protein preparation as immunogen, including GPCR immunogens, ion channel immunogens, and transporter immunogens, Magic™ platform provides the following strategies:

Cell-based Membrane Protein Expression

Creative Biolabs has developed a cell-based membrane protein expression system, which can yield membrane proteins of interest from a recombinant plasmid in living cell system. This platform is highly flexible for multiple options of host organisms, including:

• Membrane Protein Expression in E. coli
• Membrane Protein Expression in Yeast Cells
• Membrane Protein Expression in Baculovirus-Insect Cells
• Membrane Protein Expression in Mammalian Cells
• Membrane Protein Expression in Photosynthetic Bacteria

Cell-free Membrane Protein Expression

Instead of using integral cell systems, a cell-free platform which is derived from modified cellular extracts, exert unique beneficial feature for expressing many challenging or “difficult” target, especially multipass membrane proteins or toxic proteins. This highly robust, efficient and controllable system can also be directly incorporated with lipids or nanodiscs, avoiding the use of solubilizing detergent. Creative Biolabs now provides versatile cell-free expression service using multiple well-established systems:

• Cell-free Protein Expression in E. coli Lysate
• Cell-free Protein Expression in Wheat Germ
• Cell-free Protein Expression in CHO Cell Lysate
• Cell-free Protein Expression in Rabbit Reticulocyte Lysate
• Cell-free Protein Expression in Insect Cell Lysate
• Cell-free Protein Expression in HEK Cell Lysate
• Cell-free Protein Expression in PURE System

Anti-Membrane Protein Antibody Production Using Lipoparticles

We can genetically integrate the indicated membrane protein (especially viral coat proteins) on virus-like particles (VLP), a kind of self-assembling but non-infectious virus. Amplifying retroviral (HIV-1 or MLV) gag protein in 293 or CHO-based EBEE™ technique, our patent technique is able to introduce adequate post-translational modification pattern in the expressed membrane protein.

Membrane Proteins Reconstituted in Micelles

Detergent-solubilization is a routine procedure for membrane protein isolation. Selection of appropriate detergent is very critical as misuse of detergent might compromise protein structure, resulting in denatured products. Based on our specific reagents together with other commercially available detergents, we can perform up to 20 conditions to screen for the optimal solubilization outcome. As an alternative, we also offer detergent-free plan using our cell-free platform in case detergent use may cause inevitable problems.

Anti-Membrane Protein Antibody Production Using Proteoliposomes

Proteoliposomes are one of the most frequently used formats for stabilizing membrane proteins, which can be used for immunization, assays and many other applications. We have several different lipid compositions available (natural or artificial) for membrane protein reconstitution. With our special protocol and extensive experience, various multipass membrane proteins have been successfully integrated into highly-purified and ready-to-use liposomes.

Anti-Membrane Protein Antibody Production Using Nanodiscs

Nanodisc, a synthetic model that maintained by lipids and amphipathic membrane scaffold proteins (MSPs), represents a near-native phospholipid bilayer environment. This technology is quite advantageous over conventional liposomes due to its more compact lipid composition and higher stability. Nanodisc system is especially suitable for GPCR stabilization.

Anti-Membrane Protein Antibody Production Using Polymers

Creative Biolabs also offers polymer reconstitution service of membrane proteins, using amphipols and poly (styrene-co-maleic acid) lipid particles (SMALPs). Polymer technique can solubilize hydrophobic membrane proteins without detergent, meanwhile maximally preserve native confirmation and bioactivity.

Anti-Membrane Protein Antibody Production by Whole Cell Antigen Presentation

Creative Biolabs is professional in constructing membrane protein overexpressing stable cell lines. Currently, we have over 200 GPCR/ion channel stable cell lines (CHO/HEK background) with high expression levels up to 5 million per cell. We can employ these ready-to-use cell lines for membrane protein isolation or direct whole-cell immunization, bypassing time- and cost-consuming heterogeneous production.

Anti-Membrane Protein Antibody Production Using Designed Peptides

Well-designed peptide antigens derived from extracellular an intracellular loops of target proteins can raise functional antibodies that recognize the native conformation of many multi-pass integral membrane protein antigens with higher specificity, accessibility, and lower toxicity. This can be a much easier plan if the antigen domain is fully characterized.

Chimeric Soluble Proteins

Our MPAT™ Multi-spanner Antibody Technology is able to overcome the difficulty of multi-spanner membrane protein expression by replacing the transmembrane domains with a soluble peptide, thereby presenting the antigen as a chimeric soluble protein.

Anti-Membrane Antibody Production by DNA Immunization

Creative Biolabs also provides DNA immunization to discover novel anti-membrane protein antibodies. With preliminary well-characterized protein sequences, this strategy can avoid antigen preparation process and maximize the likelihood of generating and maintaining the native protein structure. It has been proved exceptionally effective in mounting strong antibody response for the challenging targets.

For anti-membrane protein antibody discovery, the Magic™ platform has developed various methods from which you can select the best fit for your specific project:

With extensive expertise and unparalleled technologies, we have gained remarkable success in developing hundreds of impeccable antibodies against diverse membrane proteins such as:

• Anti-GPCR Antibody
• Anti-Ion Channel Antibody
• Anti-Transporter Antibody

Anti-Membrane Protein Antibody Discovery by Phage Display Technology

With our unparalleled phage display immune library construction and screening platform, Creative Biolabs is professional in tailoring high specific, high-affinity antibodies targeting various membrane proteins. We provide plenty of host species choices, including rat, mouse, rabbit, guinea pig, llama, camel, sheep, horse, etc.

Anti-Membrane Protein Antibody Discovery Using Hybridoma

Hybridoma has been a long-term classic approach for monoclonal antibody production. With our proprietary techniques, we can raise hybridomas from mice, rats, hamsters, guinea pigs, etc. Of note, we utilize a special adjuvant that guarantees strong profound immune responses, which is the key for ideal antibody generation.

Anti-Membrane Protein Antibody Discovery Using Humanized Transgenic Mice

Creative Biolabs has introduced multiple best-in-class transgenic mice for generation of fully human antibodies. These genetically modified mice are capable of raising highly specific, high-affinity human antibodies after immunization, thus eliminating tedious humanization and affinity maturation process. Different genetic background and special engineering procedures can be used to ensure adequate immune responses.

Anti-Membrane Protein Antibody Discovery via Native™ Platform

Based on antigen-specific B lymphocytes cytometry technology, our Native™ Antibody Discovery Platform can screen for single antigen-specific plasma cells to retrieve native antibodies with extremely superior in vivo properties. Antibody binders directly isolated from plasma cells not only ensures native-paired V_H-V_L but also benefit from in vivo selection and affinity maturation.

Anti-Membrane Protein Antibody Discovery through Premade Antibody Libraries

Notably, Creative Biolabs carries multiple first-class premade human antibody libraries with overwhelming capacity and diversity that can derive high-quality binders against many targets. Customers can rely on our extraordinary library resources and extensive expertise in library biopanning. The premade antibody libraries available in Creative Biolabs including:

• Premade Human and Mouse Antibody Libraries
• Premade Phage Display Peptide Libraries
• Premade Single Domain Antibody Library
• Immunized Human Antibody Libraries

Equipped with world-leading technology platforms and professional scientific staff, Creative Biolabs is always dedicated to assisting our clients with the most satisfactory membrane protein antibody related services. If you are interested in discovering novel membrane protein antibodies, please feel free to contact us for more details.

Creative Biolabs is a world’s leading service provider for bio-molecular interaction analysis. Our expert team offers the most professional solutions and the advanced technologies which will ensure the progress of our customers’ research. Our expertise including but not limited to identifying, analyzing, and validating the bio-molecular interactions and also the discovery of novel drug target.

The interactions between bio-molecules play key roles in many essential biological processes. While some proteins function independently, a large number of proteins are only active in complex forms, which are defined by their interactions with other biomolecules, such as proteins, DNAs, RNAs, lipids, and metals. The bio-molecular interaction not only describes the physical contact among protein partners but also expands to the functional correlations in metabolic pathways, signaling pathways and even genetic co-regulations. Bio-molecular interactions are essential in almost all the aspects of cellular processes, such as signal transduction, regulation of metabolic pathways, environment sensing, cellular motion. Therefore, understanding the contact between bio-molecules take place in fundamental cellular processes and how bio-molecule complexes function in a cellular network becomes increasingly important.

Protein-Protein Interaction Assay

Protein-protein interactions (PPIs) as putative therapeutic targets for new treatments is challenging but gaining ground in the drug discovery industry for its significant progress in many diseases including cancer, cardiovascular disease and so on. Creative Biolabs has extensive experiences and advanced approaches on protein-protein interaction study. We could help you map the interacting network for your target proteins, validate the binding partners, and also design, discover and decipher the novel drugs.

Fig.1 Protein-protein interaction.

In Creative Biolabs, the approaches for PPI detection are mainly classified into three types, in vivo, in vitro, and in silico methods. In vivo techniques include yeast two-hybrid system (Y2H), mammalian two-hybrid system (M2H), bacterial two-hybrid system (B2H), reverse two-hybrid system (rY2H), etc. For in vitro way, Co-immunoprecipitation (Co-IP), pull-down assay, protein-fragment complementation assay (PCA), phage display, X-ray crystallography, NMR spectroscopy, and surface plasmon resonance (SPR) can be offered. And in silico groups consist of text mining and computational analyses, are displayed on a computer or by computer simulation, involving gene fusion, sequence-based approaches, chromosome proximity, and so on.

Protein-Nucleic Acid Interaction Assay

Protein-nucleic acid interactions (PNIs) are fundamental for maintaining genome and reproducing life in central biological process ranging from DNA replication and repair to RNA packaging and maturation, recombination and transcription of DNA to translation and transport of RNA. These specific interactions are susceptible to monitor through several well-established analytical modes from Creative Biolabs involving biochemical, biophysical, genetic, and immunological techniques based on the diverse purpose of our clients.

Fig.2 Protein-DNA interaction.

In Creative Biolabs, in vitro services used for PNI contain electrophoretic mobility shift assay (EMSA), chromatin immunoprecipitation (ChIP), DNA/RNA pull-down, as well as other sensitive techniques such as aptamer by SELEX, X-ray crystallography, surface plasmon resonance (SPR), and NMR spectroscopy. For in vivo assays, there are yeast one-hybrid (Y1H), yeast three-hybrid (Y3H) and fluorescent resonance energy transfer (FRET). And the identification of PNI in silico is aided by sequence-based, structure-based, or other database and bioinformatics simulation through computational intelligence.

Creative Biolabs has long-devoted in providing bio-molecular interaction analysis solutions. From experiment design to data analysis, we offer one-stop and step-to-step custom services to facilitate your project success.

Creative Biolabs has extensive experience to offer the novel human or humanized antibody services for either therapeutic or laboratory applications. We have three routes in developing human antibodies. The first and most straightforward method is to screen our human antibody libraries. The second method is to immunize premade “humanized” mice and then raise hybridoma cell lines [or to create an immune humanized antibody library] that produce human antibodies. The third method is antibody humanization. We rely on animals to produce non-human antibodies using either hybridoma or immune antibody library screening and screening methods; after that, the top candidate is humanized.

Through the flourish of antibody based therapeutic drugs, the demand of human or humanized antibody is synchronized growth. Due to the potential risk of immunogenicity, the full human antibody or at least humanized antibody (from xenogeneic sources) is required for further clinical application. Because of the serious ethical and moral issues, it is obviously an impossible task to directly immunize human beings with interested antigens. Nevertheless, several techniques have been developed as alternative solutions to overcome this limitation. Creative Biolabs is able to provide human or humanized antibodies in the ethical standards category, three main aspects have been enumerated as followed:

Truly Human Antibody from Premade Human Antibody Libraries

Creative Biolabs owns several naive or synthetic human antibody libraries in-house with diversity of over 10⁹. These premade libraries are generally based on human antibody genes from non-immunized donors or human antibody repertoire and constructed in scFv or Fab format. In this way, these premade libraries are qualified for screening human antibodies specific binding to the target protein (or other molecules). Owe to the 100% human sources, the isolated antibodies carry rare risk to cause significant immunogenicity issue in therapeutic applications.

Fully Human Antibody from FHAT Transgenic Mice

Another choice for producing fully human antibody is utilizing the Fully Human Antibody Technology™ (FHAT) of Creative Biolabs. This technology is based on the creation of a mouse strain engineered to produce a large repertoire of human antibodies in the absence of mouse antibodies. Through introducing human immunoglobulin loci into the germline of mice deficient in mouse antibody production, the “humanized” transgenic mice (FHAT transgenic mice) are able to produce significant levels of antigen-specific human antibodies with high affinity and specificity.

Non-human Primate (NHP) Antibody Humanization

Creative Biolabs has developed the innovative Hi-Affi^TM platform to provide top-class humanization of non-human primate (NHP) antibody as a unique solution to develop therapeutic antibodies for human use. Compared with the commonly used rodent, NHP has much closer genetic relationship to that of human, which means that the antibodies isolated from immunized NHPs may have higher affinity and lower immunogenicity to human beings. On the other hand, the humanization of NHP antibody is much easier than other species, and potentially has no affinity loss under suitable technique.

Besides non-human primate, Creative Biolabs also provide humanization service to antibodies derived from other species, such as mouse, rat, rabbit, chicken, llama, dog, shark and more.

Creative Biolabs provides a comprehensive antibody analysis services to make the best understanding for interested antibody property. We offer tailored solutions for antibody analysis to support drug discovery, manufacturing process and any other particular requirements.

Antibody products, monoclonal antibody (mAb) especially, have played a major role of the growing biologics drug market. For the diagnosis and treatment of various disease types, the development of monoclonal antibody therapeutics is almost the largest and most effective choice for oncological, immunological and anti-infective therapeutic areas. One reason of this interest is due to the exquisite specificity of mAbs. In general, a particular antigen can be specifically bound by a monoclonal antibody which has structurally identical antigen-binding regions recognize the same site of the antigen. This specificity can lead to highly selective interactions with interested targets involved in the pathology of disease, such as receptors, tissues and organs, and further minimizing the potential side effects.

While, the complex nature of antibody products presents a number of unique challenges during their development and manufacture of biopharmaceuticals. During the development and production of antibody based therapeutic drugs, it is essential to detect, characterize and quantify impurities as well as the important parameters about structural variants and modifications, affinity and immunogenicity to monitor the stability and safety of the final products. This physical truth has been emphasizing the need for innovative and efficient analytic tools and services. Fortunately, Creative Biolabs is just the chosen one who is able to meet clients’ analytical needs by offering a wide range of services that have been specifically tailored to meet the quality analysis and high-throughput requirements of antibody therapeutic development projects. These services include de novo antibody sequencing, affinity measurement, epitope binning, immunogenicity assessment, isothermal titration calorimetry (ITC), differential scanning calorimetry (DSC), dual-polarization interferometry (DPI) and more.

Creative Biolabs is a long-term expert who provide remarkable antibody related services. Our professional academic team have concentrated on the research and development of antibody realm for over decades and successfully contributed a range of services specifically tailored to antibody analytics. We are confident in applying our experience, industry and regulatory knowledge to design actual strategic packages pertinent to customers’ project.

Fig. 1 Antibody Basics.

Creative Biolabs is a leading service provider that focuses on antibody engineering for research, diagnostic and therapeutic use. Our service portfolio includes antibody affinity maturation, antibody stability improvement, chimeric antigen receptor (CAR) construction, scFv/Fab construction and other antibody specialization and modification services.

Since the first hybridoma has been obtained, a kind of possibility has arisen to modify the antibody to enhance its functionality. During this decade, antibody has become a major class of new drugs to treat several disease indications including cancer and autoimmune diseases. As the foundation of many diagnostics and therapeutics, the specific antigen binding ability of antibodies is critical for variety purposes, such as expanding detection limits, extending dissociation half-life time, decreasing therapeutic dosages and increasing overall efficacy. Nevertheless, it is still beneficial to optimize these antibodies or antibody related molecules with respect to their activity.

Through modern molecular biology techniques, interested antibodies can be specific modified or engineered to suit the application. If the heavy- and light-chain sequences are determined, the interested antibody can be reconfigured into different isotypes or a variety of formats (scFv, Fab, sdAb, bispecific antibody, etc.) and other possible modification including affinity measurement, humanization, caninization and so on. One feasible way is done by sequencing and cloning the interested region of antibody and grafting this select region onto a suitable framework to construct a plasmid. The predicted antibody can then be produced by transient transfection using appropriate expression system, such as mammalian cells and E. coli strains. As a choice, antibody modification can also incorporate with phage or other display methods to obtain a much broader candidates or select the most ideal products for numerous applications. Notably, all these modification and manufacture can be perfectly implemented by Creative Biolabs who is a well-recognized expert in the field of antibody development and phage display technology.

Creative Biolabs has devoted to the field of antibody engineering for over decade and established a series of comprehensive platforms to integrate various antibody engineering services. With the extensive experience, abundant industry knowledge and advanced technical skills, our scientists are fully confident in accomplishing even the most challenging antibody engineering project.

Creative Biolabs can offer various antibody labeling services to help our customers obtain their very own labeled antibodies. Our technical scientists can not only save the valuable time, but also provide highly stable and qualified products for our clients all over the world.

Labelled antibodies have become essential research tools in the biological researches as well as diagnostics. Since antibodies have been widely used to detect and quantify antigens, a large amount of demands of labeled antibody have generated. Currently, a variety kinds of labels are available to be used, such as organic dyes, fluorescent proteins, colored particles, enzymes and other potential bioactive molecules which have unique properties to produce measurable signal under the appropriate conditions. For instance, horseradish peroxidase (HRP) and alkaline phosphatase (ALP) can be attached as signal-generating molecules via the specific reaction they catalyze, while fluorescent labels are detected by microscopy equipment under excitation at specific wavelengths.

With appropriate strategy, various labels can be linked to specific antibodies. In general, four modes of labelling strategies can be used to the antibody molecules, which include linkages occur through charged amino acids, carbohydrate moieties, sulfhydryl groups and tyrosine residues. In detail, charged amino acids are abundant and wildly distributed on the outer surface of antibodies, however the modification may interfere with the function of antibody; carbohydrate moieties are generally found on the highly glycosylated Fc region which are distant from the antigen recognition site, while required more attention to the oxidation steps; sulfhydryl groups contains on cysteine residues, the modification may not damage the specificity but may affect the avidity; tyrosine residues can be specifically iodinated as radioactive label, although has fallen out of favor. Due to the different advantages and shortcomings, choosing or combining suitable strategies are critical to build up an ideal labeled antibody. Because of the ability to image multiple antigens in a single cell or tissue section, labelled antibodies has allowed many elegant and valuable experiments to be performed and expanded their applications to a variety of immunoassays, such as flow cytometry, ELISA, western blotting and immunohistochemistry. Nowadays, Creative Biolabs is able to provide novel labelled antibodies suitable for customers’ particular application needs.

As an acknowledged biotechnology provider, Creative Biolabs is noted for its first-class services, strictly controlled products and customer oriented philosophy. By leveraging years of antibody labeling expertise, our scientists are able to provide a wide selection of services for both research and commercial use with the best quality at the most competitive price.

Features of the Services:

• A full range of services from protein purification and labeling to conjugate purification.
• A wide variety of labels including fluorescent dyes, horseradish peroxidase, alkaline phosphatase and other enzymes, biotins and haptens.
• Instant access to protein expression and antibody production services.
• The most competitive prices in the industry.

Creative Biolabs offers a full spectrum of bioconjugation services, including protein/antibody bioconjugation, oligosaccharide bioconjugation, oligonucleotide bioconjugation, surface immobilization, PEGylation, synthetic polymer modification and liposome conjugation services. Additionally, we also offer custom bioconjugation service to meet your very unique research objectives.

As a leading service provider that focuses on novel therapeutics and medicine development, for over a decade, Creative Biolabs has been constantly delivering numerous hybrid complexes that are supported by our state-of-art technical platform and rich expertise. From project scope analysis and problem solving proposals, to complex biopolymer production and assay development, we succeed with excellence and confidence to meet your demand for creating novel bioconjugates, which are efficient and cost-effectively.

Our featured services contain, but are not limited to:
• Antibody-dye conjugation
• Antibody-enzyme conjugation
• Antibody-drug conjugation
• Antibody-biotinylation
• Fab PEGylation
• Streptavidin-biotin system
• Bioconjugation for protein-protein interaction study
• Vaccines and immunogen conjugation
• Isotope labeling
• Gold nanoparticle labeling
• Polystyrene bead labeling

Bioconjugation involves the attachment of one biomolecule to one or several other molecules, usually through a covalent bond. There are endless number of applications using bioconjugates, such as in purification, capturing, scavenging, catalysis, tracking and imaging, as well as diagnostic and therapeutics areas, assay development, biomolecule quantification and detection, vaccine and drug design. Our comprehensive and proficient bioconjugation technical platform can deliver successfully, specific, efficient bioconjugations services to meet you very specific requirement in research objective, budget and time.

To design a successful bioconjugates while still maintaining all the functions of its individual parts, it requires sufficient knowledge of the reactive groups, reagent types, and functional groups present on the biomolecules, as well as an understanding of how modification of a biomolecule can potentially affect its activity. Our Ph.D level experts have rich knowledge and extensive experience in bioconjugation chemistry. Our scientists are ready to assist you step by step from the bioconjugation design, strategy determination to the final implementation and manufacturing to fit your specific need.

Choose our bioconjugation services by molecule:

• Protein/Antibody Bioconjugation
• Biotinylation
• Oligosaccharides Bioconjugation
• Oligonucleotide Bioconjugation
• Surface Immobilization
• Liposome Conjugation
• PEGylation
• Synthetic Polymer modification
• Featured Conjugation Services

Need help on how to select? Our Ph.D level scientists are ready to be of your assistance.

Creative Biolabs offers comprehensive immunologic analysis services for a wide range of biotherapeutic drug candidates, including protein, enzyme, antibody, an antibody-drug conjugate (ADC) and so on. Based on our experienced scientist team and elaborate technical platforms, we are capable of carrying out your assay with optimized and validated protocols in a fast and reliable manner.

Remarkable characteristics of prolonged half-life, high target specificity, and low intrinsic toxicity make biotherapeutic drugs significantly outperform small molecule drugs, and attract increasing attention in the global academic and pharmaceutical industrial field. However, the use of biotherapeutic drugs may lead to unwanted immune responses, resulting in the efficacy decrease of the drugs and even safety threaten to patients. It is worthwhile and necessary to conduct complete immunologic analysis for biotherapeutic drug candidates and biosimilar biologics at the early stage of drug development. By taking this into account, Creative Biolabs integrates four delicate platforms together to provide high-quality services packages to help clients evaluate their drug candidates, and the contents of our immunologic analysis services are listed below.

SIAT® Immunogenicity Assessment Services

The immune responses to biotherapeutic drugs can decrease the efficacy of the drugs due to faster clearance of the drugs as immune system treat them as antigens and launch specific immune reaction. Furthermore, unwanted immune system activation may cause rapid and enormous cytokine release and bring safety to threaten to patients. Therefore, it is of great importance to conduct Immunogenicity risk management of biotherapeutic drugs and biosimilar biologics in the preclinical stage. Creative Biolabs provides the proprietary Sensitive Immunogenicity Assessment Technology® (SIAT®) system to help clients evaluate the immunogenicity potential for a broad range of biotherapeutic drug candidates, and screen out optimal individuals.

Fig.1 Immunologic Analysis Services.

ECIA™ Cellular Analysis Services

As the ancillary service of SIAT® Immunogenicity System, Creative Biolabs offers ECIA™ cellular analysis services designed to accelerate your immune monitoring or epitope discovery projects in a cost-efficient way. Our customized ECIA™ cellular analysis platform can be a good choice for clients who are eager to acquire HLA allele information, to identify antigen-specific B cells and T cells, to detect Intracellular cytokine level as well as to monitor specific T cell proliferation. Based on our unique high-throughput technique, enriched experience, and professional service team, Creative Biolabs promises to provide the most suitable cellular assay design and deliver accurate reports with reliable data for each project.

CreMap™ Epitope Mapping & Discovery Services

Creative Biolabs has developed an excellent CreMap™ epitope mapping & discovery platform to help customers identify the specific binding sites of antibodies (or BCRs, TCRs) on target antigens, and those binding sites on antigens are so-called epitopes. The process of identifying epitopes of antibodies (or BCRs, TCRs) on certain antigen is known as epitope mapping, which plays a crucial role in the discovery of new biotherapeutic drugs, vaccines, diagnostic reagents, and so on. Our CreMap™ platform offers both B cell epitope mapping and T cell epitope mapping service to assist your research or drug discovery project in a cost-efficient way.

RDIT® De-immunization Services

Eliminating the immunogenicity of therapeutic proteins or biotherapeutic drugs is critical to improving clinical outcomes. Creative Biolabs has developed a well-mature RDIT® de-immunization platform to offer a number of assays to diminish the immunogenicity. In general, de-immunization can be accomplished by many strategies, such as amino acid modification, T/B cell epitope identification, as well as structure reconstruction. In addition, we have also generated a computer-based tool to produce non-immunogenic versions of potential targets. This tool can determine the immunogenic region of biotherapeutic molecules to identify specific analogs with low binding affinity.

Service Highlight

• Efficient and accurate immunogenicity evaluation
• Broad population coverage
• Fast and cost-effective evaluation
• A range of protein engineering capabilities

Creative Biolabs’ technical experts will help you choose the best service to meet your specific needs. For more details on how we can provide innovative support to your project, please feel free to contact us.

Manufacturability assessment offers the opportunity to evaluate the technical viability of a drug candidate at the industrial scale and functions as a bridge between discovery and process development stage. Creative Biolabs is able to evaluate key manufacturability parameters, including productivity and process yield, chemical stability, PTMs, physical stability, aggregation, electrostatic properties, and formulability through our virtual prediction platform and experimental manufacturability testing platform. Pilot studies conducted by our labs have suggested that these results can effectively guide the customer to the most appropriate candidate for process development. Meanwhile, we have also developed a variety of assays to improve the developability condition of potential drugs or protein therapeutics.

Introduction to Manufacturability Assessment and Optimization

Peptides, antibodies, or proteins have shown promising results in treating many kinds of diseases, but side effects still happen and are usually related to the physicochemical property of testing candidates. The safety and efficacy of candidate drugs still need to be analyzed by “manufacturability”. Manufacturability assessment and optimization play an important role in detecting the suitability of a therapeutic molecule. As a result, recent studies have revealed that high-throughput methods should be a powerful tool for eliminating the instabilities, toxicity in successful manufacturability projects. In general, comprehensive assessment and optimization are based on computational and experimental analysis strategies. In silico prediction, technologies have been widely used for modifying the structure and sequences of screened molecules to improve their stability and manufacturability in clinical use. In addition, the application of experimental approaches is an emerging area, and much progress has been made to detect the solution properties, chemical stability at different concentrations.

Fig.1 Manufacturability Assessment and Optimization Platform.

Virtual Prediction of Manufacturability Assessment and Improvement

As an expert in manufacturability assessment, Creative Biolabs has generated a novel in silico analysis platform to provide a series of services to measure multiple manufacturability parameters in diverse target molecules. This platform can rapidly evaluate physical and chemical parameters, including viscosity, solubility, critical residues, aggregation, immunogenicity as well as degradation/ PTMs, in a cost-effective manner. Furthermore, we also offer a suite of predictive models for engineering the structure of protein therapeutic candidates to improve the evaluation results of manufacturability. For instance, The aggregation and stability risks of monoclonal antibodies against CD19 have been assessed by our in silico tools.

Experimental Manufacturability Assessment and Optimization

In recent years, Creative Biolabs has designed a novel experimental platform to screen and optimize the manufacturability of test materials in drug research. Up to now, we have successfully accomplished a wide collection of early assessment strategies that can be used to analyze physicochemical stability parameters, deamidation, aggregation propensity, oxidation propensity, as well as solution properties. Subsequent screens and assays are focused on binding specificity and affinity specificity. A number of high-throughput (HT) technologies, such as fluorescence spectroscopy, Nuclear magnetic resonance (NMR), high-performance liquid chromatography (HPLC), dynamic light scattering (DLS), have been proved its applicability in the screening cascade. Additionally, our customize manufacturability optimization services are designed to provide meaningful data for designing better candidates in drug discovery.

Fig.2 Manufacturability assessment of the drug candidates.

Creative Biolabs offers a manufacturability assessment and optimization platform that is critical to improving the quality, safety, and manufacturability of potential biotherapeutic and drug candidates. If you are interested in our service, please feel free to contact us.

Creative Biolabs has integrated multiple cutting-edge technologies and established a powerful Magic™ membrane protein antibody platform. In addition to antibody generation, our scientists are also specialized in producing high-quality membrane protein products (both membrane crude or purified proteins) for immunogen preparation, assay development, and various other applications.

Membrane proteins, which constitute over thirty percent of all proteins in human genome, play a vital role in many cellular functions like materials transportation, signal transduction, intercellular recognition ligand-receptor binding, cell adhesion, etc. As such, they represent the absolute majority (over 60%) of current drug targets. Due to their low natural abundance and highly complicated structure (partially or completely associated with plasma membrane), to obtain recombinant membrane protein products with native confirmation and bio-activity has always been a challenging issue.

During the past decade, scientists in Creative Biolabs have devoted to various optimizations and exploration to process development of membrane protein expression, solubilization, purification, and characterization. Our versatile Magic™ membrane protein platform employs diverse state-of-the-art techniques to pursue the best-quality membrane protein products With years of experience, we have gained remarkable success in obtaining a variety of difficult membrane proteins such as G-protein coupled receptors (GPCRs), ion channels, membrane transport proteins, etc.

To facilitate your antibody discovery and development against various membrane proteins, Creative Biolabs has developed a serial of membrane proteins with multiple species and tags. The proteins have been identified to be with high purity and low endotoxin. The protocol for bioactivity assays can be provided for free upon request. Please browse our membrane proteins here.

Magic™ membrane protein platform offers extremely flexible options for expression, solubilization, reconstitution, and characterization, including but not limited to:

Cell-based Membrane Protein Expression

The cell-based heterogeneous expression is the most commonly used approach for recombinant proteins. For membrane proteins, it can be problematic as for adequate expression levels, correct membrane insertion, and native confirmation preservation. Using our proprietary vector designs and special protocols, we have successfully generated a broad spectrum of targets. Our impeccable platforms can provide multiple options of host organisms, including:

• Membrane Protein Expression in E. coli
• Membrane Protein Expression in Yeast Cells
• Membrane Protein Expression in Baculovirus-Insect Cells
• Membrane Protein Expression in Mammalian Cells
• Membrane Protein Expression in Photosynthetic Bacteria

Cell-free Membrane Protein Expression

Instead of using integral cell systems, a cell-free platform which is derived from modified cellular extracts, exert unique beneficial feature for expressing many challenging or “difficult” target, especially multipass membrane proteins or toxic proteins. This highly robust, efficient and controllable system can also be directly reconstituted with lipids or nanodiscs, avoiding the use of detergent. Creative Biolabs now provides versatile cell-free expression service using following systems:

• Cell-free Protein Expression in E. coli Lysate
• Cell-free Protein Expression in Wheat Germ
• Cell-free Protein Expression in CHO Cell Lysate
• Cell-free Protein Expression in Rabbit Reticulocyte Lysate
• Cell-free Protein Expression in Insect Cell Lysate
• Cell-free Protein Expression in HEK Cell Lysate
• Cell-free Protein Expression in PURE System

Virus-like Particles (VLPs) and Lipoparticles

Virus-like particles (VLPs) are multiprotein structures that mimic the organization and conformation of authentic native viruses but lack the viral genome (non-infectious). They are widely applied in vaccinology, immunology, and virology. Currently, we offer VLP construction and purification service in various systems, including bacterial, insect, mammalian, etc. PTMs can also be incorporated.

Membrane Protein Stable Cell Lines

Notably, except recombinant expression, Creative Biolabs is also professional in constructing membrane protein overexpressing stable cell lines. Currently, we have over 200 GPCR/ion channel stable cell lines (CHO/HEK background) with high expression levels up to 5 million per cell. These elegant cell lines can be directly used for membrane isolation, immunization and various assays (like SPR), which significantly saves valuable research time.

Membrane Proteins Reconstituted in Detergents

Detergent-solubilization is a routine procedure for membrane protein isolation in most cases. Selection of appropriate detergent is very crucial as misuse of detergent might compromise protein structure, resulting in denatured products. With our unique reagents as well as other commercially available detergents, we can perform up to 20 solubilizing conditions to screen for the best outcome. As an alternative, we also offer detergent-free plan using our cell-free platform.

Membrane Proteins Reconstituted in Liposomes

Proteoliposomes are one of the most frequently used formats for stabilizing membrane proteins, which is well applicable to immunization, assays, and diverse usages. We have several different lipid compositions available (natural or artificial) for membrane protein reconstitution. Aided by our well-established protocol and vast experiment, the protein products can be incorporated into lipids with correct folding and high bio-activity.

Membrane Proteins Reconstituted in Nanodiscs

Similar to proteoliposomes, nanodiscs also aims at stabilizing membrane proteins in mimic lipid bilayer, yet with a more dense environment (formed by membrane scaffold proteins, aka MSPs) that is even closer to natural membrane environment. Therefore, nanodisc technology has several advantages over conventional methods, especially for multipass membrane targets like GPCRs, ion channels, etc.

Membrane Protein Production Production Using Polymers

Creative Biolabs also offers polymer reconstitution service of membrane proteins, using amphipols and poly (styrene-co-maleic acid) lipid particles (SMALPs). Polymer technique can solubilize hydrophobic membrane proteins without detergent, meanwhile maximally preserve native confirmation and bioactivity.

In addition to intact/ full-length membrane proteins, we are also specialized in expressing recombinant peptides/fractions derived from extracellular or intracellular loops. This is particularly suitable for many large, multi-spanner membrane proteins with well-characterized structure. Compared with full-length expression, it will be much easier to obtain recombinant peptide domains of interest, which can also work perfectly in immunization, antibody selection, etc. Moreover, we can also replace the transmembrane domain with a soluble peptide, creating a chimeric soluble protein with indicated extracellular/intracellular domains.

Equipped with multiple leading technologies and abundance expertise, Creative Biolabs is confident in delivering satisfactory membrane protein products with our first-class professional service. We provide quite flexible options about expression, solubilization/reconstitution and purification as well as characterization (such as binding assays, functional assays). A pilot study is strongly recommended to gather some preliminary date, and then we can move to further optimizations and scaling-up stage. Please feel free to contact us for a detailed quote.

Creative Biolabs provides professional custom services in membrane protein characterization to facilitate our customers’ research, development, and applications of membrane proteins. Our versatile platform is compatible with various structure, kinetics and function profiling of membrane proteins. This service can be offered separately or as an integral step of our membrane protein preparation package, to ensure that the final product can meet custom standards of purity, function, structure, binding behaviors, etc.

Our membrane protein characterization services including but not limited to:

• Native polyacrylamide gel electrophoresis (PAGE)
• SDS polyacrylamide gel electrophoresis (PAGE)
• Fluorescence spectroscopy
• Circular dichroism spectroscopy
• UV-Vis absorption spectroscopy
• Chemiluminescence
• Radiometry
• Dynamic light scattering (DLS)
• Analytical gel-filtration/size-exclusion chromatography
• Analytical ultracentrifugation
• Western blotting
• Enzyme-linked immunosorbent assay (ELISA)
• Immunoprecipitation
• Surface plasmon resonance (SPR) and bio-layer interferometry (BLI)
• X-ray crystallography of membrane proteins
• Nuclear magnetic resonance (NMR) spectroscopy
• Cryo-electron microscopy (Cryo-EM) of membrane proteins
• SAXS studies
• Function validation

From these services, our customers can obtain information for membrane protein structure, function, activity as follows:

• Purity and composition
• Topology, size, and oligomeric state
• Atomic structure
• Thermodynamic and kinetic parameters for binding with drugs, antibodies, and ligands
• Thermodynamic and kinetic parameters for folding and oligomerization
• Enzyme activity
• Signal transduction efficiency and selectivity
• Molecule and ion transportation efficiency and selectivity

Scientists in Creative Biolabs have decades of experience and abundant knowledge in membrane protein characterization. We will be happy to explore the world of membrane proteins together with our customers. Please feel free to inquire us for more information.