Bispecific Antibody Analysis Service

Alpha Lifetech can provide a series of bispecific antibody purification, antibody sequencing, antibody validation and other services. We can provide our customers with quality assured antibody and recombinant protein products and services. We can prepare antibodies with high efficacy, strong specificity, and good stability.

Bispecific antibodies are a type of genetically engineered antibody, mainly prepared by cell fusion or recombinant DNA. Bispecific antibodies have two specific antigen binding sites, allowing them to specifically bind to two antigens or two different epitopes on the same antigen simultaneously.

Advantages of Bispecific Antibodies

Compared with monoclonal antibodies, bispecific antibodies have two antigen binding sites and can simultaneously bind to two different specific epitopes or target proteins. Therefore, bispecific antibodies have stronger specificity and can more accurately target target target cells or molecules. Bispecific antibodies can enable immune cells such as T cells and NK cells to form overseas connections with tumor cells. By recruiting immune cells around tumor cells and activating them, the killing power of bispecific antibodies against tumors can be enhanced. At the same time, bispecific antibodies can also block two different signaling pathways, thereby enhancing cell killing effects. After binding to two different cell surface antigens, the binding specificity of bispecific antibodies may be enhanced, and the occurrence of off target side effects may be reduced. Therefore, the safety of treatment will be improved.

Based on the above advantages, the therapeutic effect of bispecific antibodies can reach 100-1000 times that of ordinary antibodies. Therefore, when using bispecific antibodies for treatment, the dosage can be significantly reduced, thereby reducing the cost of drug treatment. Bispecific antibodies also have strong competitiveness in terms of tissue permeability, tumor cell killing efficiency, off target rate, and clinical indications, and can be applied to the treatment of various diseases, such as tumors and autoimmune diseases. The structure of bispecific antibodies includes Fc based BsAbs and fragment based BsAbs. These different types of bispecific antibodies have different characteristics and advantages, and can be selected and designed according to specific needs. Therefore, the flexibility in designing bispecific antibodies is also relatively high.

Technical Difficulties

In the process of designing bispecific antibody structures, it is important to effectively combine two different antigen recognition sites while maximizing the uniformity and yield of the target antibody. In the early evolutionary process, bispecific antibodies prepared by chemical coupling or cell fusion methods had a "chain exchange problem", which means that random combinations may produce multiple different antibody structures, making it difficult to isolate the target combination.

The development of bispecific antibodies requires high technical platforms, and different technical platforms also have differences in structural design, production processes, and drug properties. There are already dozens of technology platforms for the development of bispecific antibodies, but they still need to be continuously optimized to develop platform technologies that combine drug properties, production process feasibility, and scalability.

The use of bispecific antibodies requires preclinical evaluation, and an appropriate preclinical evaluation model should be selected. The current model can to some extent reflect the mechanism of action and pharmacological effects of bispecific antibodies, but due to species differences, preclinical data is often difficult to directly convert into clinical doses, increasing the risk of clinical research.

The production of bispecific antibodies is usually low, and the purification process is complex, making large-scale preparation difficult. In the development process of bispecific antibodies, it is necessary to balance and coordinate the safety and effectiveness of two targets, ensuring that bispecific antibodies can effectively kill tumor cells and reduce side effects in clinical applications.

Fig. 1 Schematic representations of the formats and modes of action of bsAb molecules. (Figure source: Antibody-drug conjugates and bispecific antibodies targeting cancers: applications of click chemistry - PubMed (nih.gov))

production of bispecific antibodies

Bispecific antibodies can be produced through chemical conjugation, hybridoma, and genetic engineering. The technical principle is based on genetic engineering technology or cell fusion technology, which recombines gene fragments of two different specific antibodies to form antibody molecules with bispecific binding ability. This antibody molecule can simultaneously bind to two target antigens. The chemical coupling method couples two complete IgG or two F (ab ') 2 antibody fragments into bispecific antibodies by using chemical coupling agents such as phthalimide and dithiobenzoic acid. This method is simple and easy to operate, but it may damage the antigen binding site and reduce antibody activity. The four source hybridoma method can fuse somatic cells from two different hybridoma cell lines to express the corresponding mouse IgG.

Through genetic engineering technology, antibodies can be genetically modified to form bispecific antibodies. Two different monoclonal antibodies were constructed, and the Fab fragments or heavy chain and light chain variable regions of the two antibodies were cleaved separately. Through cross-linking reaction or chain recombination technology, the two fragments were combined to form a bispecific antibody. Although the complexity of genetic engineering technology is relatively high, it is currently the most commonly used method for bispecific antibody production to adjust the structure and function of antibodies. When conducting bispecific antibody design, the principle of antibody cross reactivity may be utilized, but antibody cross reactivity may cause non-specific reactions, so it will be carefully considered in the practical application of bispecific antibody design.

Bispecific Antibodies Analysis Service Workflow

Steps	Service Content	Timeline
Gene Synthesis	Generation of up to 3 different design schemes of sequences, de novo generation of recombinant antibody DNA plasmids	2-3 weeks
Small Scale Test	Production of bispecific antibody proteins → Small scale expression in mammalian cell lines → Validation by SDS-PAGE → Binding analysis with recombinant protein antigens by ELISA → Comparison of clones with parental antibodies	5-6 weeks
Identify	Expression of the first two full-length bispecific antibodies → Binding analysis by ELISA → ELISA evaluation of antibodies recognized by anti mouse antibodies	1 week
Antibody Production	Large scale antibody production	3-4 weeks

Our Advantages

We cover the upstream and downstream services of antibody production, and can provide technical services from antibody preparation, bispecific antibody purification and antibody separation and purification, antibody sequencing, antibody validation, etc, , which can be used for bispecific antibody therapy.

High Expression

High protein expression in CHO cells.
Stable Storage

>2 weeks in serum at 37 °C.
No Solubility Issue

>30 mg/ml
Stable Cell Lnes

Many stable cell lines with tumor antigens have been developed to validate many bispecific antibodies