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Antibody Modification Bioconjugation

Antibody Drug Therapy (ADC) Through Bioconjugation

Antibody conjugates have become a major highlight among biological agents for targeted therapeutic studies, and immunoassay or detection applications. Bio-Synthesis offers custom antibody bioconjugation services using monoclonal antibody or polyclonal antibody through chemical conjugation with other biomolecules. This includes the conjugation of antibodies and their fragments with biologically active molecules such as nucleic acids, oligonucleotides and their analogs, peptides, proteins, or with any molecular group that adds valuable properties—i.e. radionuclides, drugs, toxins, enzymes, metal chelates, fluorophores, haptens, and others.

Our unique and unrivaled expertise in research and development offers you the best quality antibody bioconjugates in the industry. From custom antibody development to modification and bioconjugation, all are produced in-house at Bio-Synthesis. Each antibody-biopolymer conjugate is meticulously monitored according to our stringent quality assurance and quality control standards.

Antibody Drug Therapy (ADC) Through Bioconjugation

The immunoconjugate for therapeutic purpose is undergoing an outburst of activities for research and development. This novel approach to drug delivery could lead to safer methods of reducing dosage, and increasing efficacy without causing harmful side effects. Monoclonal antibodies can be utilized for drug or isotope deliveries, however, the biological functions of monoclonal antibodies are often sensitive to subtle variations in structure. Hence, the chemical aspects of preparing, purifying, and characterizing these conjugates must be carefully considered.

Bio-Synthesis’ custom antibody conjugation services include experimental design and method development for the preparation of conjugates to be used in life science research, diagnostic and drug discovery.

Over the decades, we have accumulated a continually growing portfolio of antibody modification and conjugation services. If you can't find what you want, contact our National Customer Service Center at 800.220.0627, or contact us online with your detailed project descriptions.

Service Portfolios

  • Enzyme labeled antibodies
  • Conjugation of toxin to antibody
  • Antibody to antibody conjugation
  • Coupling of metal chelates to antibodies
  • Antibody-dye labeling
  • Antibody-DNA conjugates
  • Antibody-oligo analog conjugates
  • Antibody Luminex bead conjugation
  • Antibody-drug conjugates
  • Protein-antibody conjugates
  • Antibody-support immobilization
  • Antibody-nanoparticle conjugation
  • Antibody-macroparticle conjugation
  • Antibody-PEG or dextran modification

Sample Submission Requirements:

Modification or conjugation of whole immunoglobulin fraction, secondary antibodies, polyclonal and monoclonal antibodies should be affinity purified prior to undergoing bioconjugation to prevent nonspecific interaction. Bio-Synthesis also assists in providing affinity chromatography services using the corresponding immobilized antigen or immunoglobulin binding protein (e.g. protein A may be employed.) Client must supply Bio-Synthesis an antibody quality check documentation.

Antibody Bioconjugation Service Descriptions

Chemistry:

While traditional antibody conjugation takes advantage of lysine ϵ-amino , N-terminal α-amine groups or carboxylate groups due to its abundance in antibody, the conjugation procedure that utilize these groups often changes the overall pH of the antibody and leads to higher nonspecific binding. In addition, this random cross-linking within the antibody often obscuring the binding sites may produce partially active or inactive antibody conjugates that may not bind to the antigen. Bio-Synthesis offers site-specific antibody conjugation services based on the unique conjugation strategy developed at Bio-Synthesis. Drugs, toxins, oligo DNA, RNA, or peptide, small molecules can be conjugated to an antibody at a specific site away from the antigen binding site. Site-specific cross-linking results in high retention antigen binding activity within the ensuing conjugate. Single or few predictable sites labeling allow better manufacturing reproducibility specific binding (efficient and safe drugs).

Available Methods:

NHS Ester-Maleimide-Mediated Conjugates

Heterobifunctional reagents containing an amine-reactive NHS ester on one end and a sulfhydryl-reactive maleimide group on the other end are generally used. We provide:

  • Activate target molecule with NHS Ester-Maleimide Crosslinkers
  • Conjugate with reduced antibodies
  • Conjugate with 2-imminothiolane-modified antibodies
  • Conjugate with SATA-modified antibodies

Reductive Amination-Mediated Conjugation

Oxidation of polysaccharides residues in glycoproteins with sodium periodate provides an efficient way of generating reactive aldehyde groups for subsequence conjugation with amine or hydrazide-containing molecule via reductive amination. Many immunoglobulin molecules such as polyclonal IgG molecules often contains carbohydrate predominantly in Fc portion, they can be periodate-oxidized to contain reactive aldehyde residues to be conjugate with target molecule containing Amine or hydrazide derivatives. This is an excellent method for directing the antibody modification reaction away from the antigen binding sites if the antibody glycosylation points are solely in the FC region or way from the binding site. This method often results in the highest retention of antigen binding activity within the ensuing conjugates. We provide:

  • Activation of target molecule with sodium periodate or,
  • Activation of antibodies with sodium periodate
  • Conjugation of periodate-oxidized biopolymer to antibodies by reductive amination or,
  • Conjugation of periodate-oxidized antibodies with amine or hydrazide derivatives

Conjugation Using Antibody Fragments

It is often advantageous to use antibody fragments in the preparation of antibody bioconjugates. This is done by selective antibody fragmentation using enzymatic digestion of intact immunoglobulins into lower molecular weight molecules that still able to recognize and bind antigen. Conjugation of antibody fragment with enzyme molecules can result in ELISA reagents that possess better characteristics than corresponding conjugates prepared with intact antibody. Such antibody fragment conjugates display less interference with various Fc binding proteins and also less immunogenicity, more facile membrane penetration for immunohistochemical staining techniques, and lower nonspecific binding to surfaces or membranes, resulting in increased signal-to-noise ratios. We provide services such as:

  • F(ab')2 fragment preparation or,
  • Fab fragment preparation or,
  • Antibody reduction to two half-antibody
  • Conjugation through various crosslinking and conjugation chemistries described above.

Service Specification:

Starting quantity: 5 mgs, 2 mgs of antibody for initial pilot conjugation prior production is required for a thorough optimization and method development of purification and analysis. Although, we have been successful with as little as 200 micrograms in some cases. We provide a few test run for customer to test. Once the customer has agreed on the test conjugated within one week, Bio-Synthesis will proceed to scale up the production. After conjugation, standard desalting or purification of a small percent of heterogeneous products containing single or multi-site conjugate per molecule.

Material:

Antibody and antibody fragments: Fab, F(ab')2, Half antibody, FC,

Procedure:

All custom synthesis of biomolecule, modification or bioconjugation services is manufactured under strict quality control processes. Analytical HPLC and MS analyses are performed in every development cycle. Final target conjugates must first be isolated from excess or unreacted reagent. In many cases, simple dialysis removes unreacted reagent from the reaction solution (if the protein/antibody is significantly larger (>3-fold) than the modifying or coupling reagent). Additional purification such as stirred cell filtration, tangential flow filtration (TFF), or gel filtration chromatography may also be used to either remove excess reagent or to isolate and characterize the cross-linked product. Reagents that are similar in size or larger than the antibody (mostly protein and other biological molecules) may require other purification techniques such as affinity chromatography, ion-exchange chromatography, or hydrophobic interaction chromatography.

Cross-linked target molecule may then be further characterized by biochemical or biophysical techniques with an additional fee. Once the product has been purified, it may be subject to many different types of studies including spectroscopic (MALDI-TOF, ESI, LC-MS Fluorescence), electrophoresis, immunochemical biochemical or enzymatical. QC (quality control) and QA (quality assurance) procedures are also followed independently to offer you double guarantee for the highest quality possible of every delivered conjugates. Moreover, our dedicated technical account managers will guide your project through every step of the process and constantly keep you informed of the latest project progress.

Deliverable:

7-10 weeks, conjugates will be delivered in buffer solution close to physiological pH, such as phosphate buffered saline (PBS), which is generally recommended.

Disclaimer:

Whereas we can chemically link the intended components of a conjugated molecule, there exist the possibility that the binding sites/active sites of the protein can be altered/modified, partially or completely, independent of the stoichiometry used. Sometimes this activity loss is caused by physically blocking the antigen binding sites during conjugation or by conformational changes in the complement-determining regions. Some proteins/antibodies are just too labile to undergo chemical modification reactions, regardless of the coupling methods used.

Bio-Synthesis can only guarantee the structure of our conjugates but not the suitability to specific biological applications

If you can't find the antibody conjugation service you need, please contact us online with your detail project descriptions.

Price:

Price varies based on the project specifications. Our service includes materials and labor for conjugation only! Price does not include the cost of biopolymer synthesis or purchase from a commercial vendor and, if deemed necessary, biopolymer modification to introduce additional functional groups, extra linkers, spacers. Please contact us for a quote.

Basic Biopolymer Modification BIOCON10000
Fab Preparation BIOCON12000
Fab (ab')2 Preparation BIOCON12001
Introduction of Spacer and Linkers BIOCON12002
Introduction of Aldehyde Group BIOCON12003

Ordering and Submitting Requests for Bioconjugation Services

For us to better understand your customized project, please complete our Bioconjugation Service Questionnaire. The more our chemists understand your project needs, the more accurate feedback we will be able to provide you.  Provide us with your project details will enable to us to recommend the best reagents to use for your project.  The most useful and readily available tools for bioconjugation projects are cross-linking reagents. A large number of cross-linkers, also known as bifunctional reagents, have been developed.  There are several ways to classify the cross-linkers, such as the type of reactive group, hydrophobicity or hydrophilicity, and the length of the spacer between reactive groups.  Other factors to consider are whether the two reactive groups are the same or different (for example, heterobifunctional or homobifunctional reagents), whether the spacer is cleavable, and whether the reagents are membrane permeable or impermeable.  The most accessible and abundant reactive groups in proteins are the ϵ-amino groups of lysine.  Therefore, a large number of the most common cross-linkers are amino selective reagents, such as imidoesters, sulfo-N-hydroxysuccinimide esters, and N-hydroxysuccinimide esters.  Due to the high reactivity of the thiol group with N-ethylmaleimide, iodoacetate and a-halocarbonyl compounds, new cross-linkers have been developed that contain maleimide and a-carbonyl moieties.  Usually, N-alkylmaleimides aremore stable than their N-aryl counterparts.

In addition to the reactive groups on the cross-linkers, a wide variety of connectors and spacer arms have also been developed.  The nature and length of the spacer arm play an important role in the functionality.  Longer spacer arms are generally more effective when coupling large proteins or those with sterically protected reactive side-chains.  Other important considerations are the hydrophobicity, hydrophilicity, and the conformational flexibility.  Long aliphatic chains generally fold on themselves when in an aqueous environment, which makes the actual distance spanned by such linker arms less than expected.  Instead, spacers that contain more rigid structures (for example, aromatic groups or cycloalkanes) should be used.  These structures, however, tend to be very hydrophobic which could significantly decrease the solubility of the modified molecules or even modify some of their properties.  In such cases, it is recommended to choose a spacer that contains an alkylether (PEO) chain.  Bio-Synthesis offers several cross-linkers with PEO chains, such as thiol-binding homobifunctional reagents, heterobifunctional based, and their derivatives.

Within 3-5 days upon receiving your project scope, we will provide you an appropriate quotation. An order can be placed with PO (Purchase Order) or major credit cards ( ). Your credit card will be billed under Bio-Synthesis, Inc.