Naïve Fab Library Construction Service

Introduction of Naïve Fab Library Construction ServiceFab

Phage display technology is an in vitro screening technique that can select target antibodies from a large library of antibodies and display them on the surface of bacteriophages. The M13 phage display system, which consists of five capsid proteins, is currently widely used for display. The main function of capsid protein P Ⅲ is screening, and the target gene can be inserted into P Ⅲ, and its applicable vector range is also very wide. In 1985, phage display technology was created as a technique for studying antigen epitopes. Phage display mainly involves inserting exogenous genes into phage capsid proteins through genetic engineering technology, so that the protein encoded by the exogenous gene can be displayed as a fusion protein without changing its structure and activity. Then, through multiple rounds of screening, highly specific antibodies that can bind to the target molecule are selected. After years of development, phage display technology has been applied not only in the study of antigen epitopes, but also in areas such as cell signal transduction, protein recognition sites, and drug development. With the widespread use of phage display technology, many diseases are identified through phage display technology, which has the advantages of speed, relatively low cost, and large library capacity. By screening bacteriophage peptides that bind to specific antigens, new antibodies can be developed or existing antibodies can be improved.

Fab fragments, also known as antigen binding fragments, are key regions in antibody structures that can bind to antigens. Fab fragments are composed of a portion of the antibody's light and heavy chains, which are connected by disulfide bonds. The light chain contains a variable region (VL) and a constant region (CL), while the heavy chain contains a variable region (VH) and a portion of the constant region (CH1). VL and VH form antigen binding sites (CDRs) through hydrophobic interactions and hydrogen bonding. The molecular weight of Fab fragments is relatively small, typically between 47-55 kDa, which gives them good tissue penetration and low immunogenicity. Fab fragments retain their complete antigen binding ability and can bind with specific antigens with high affinity. Due to the lack of Fc segments in Fab fragments, immune reactions mediated by Fc segments are not triggered, and the probability of hypersensitivity reactions is also reduced. The smaller molecular weight allows Fab fragments to more easily penetrate tissue barriers and reach target sites. The half-life of Fab fragments in vivo is relatively short, which helps to reduce potential side effects of drugs. Fab antibody production can be achieved through enzymatic hydrolysis, genetic engineering techniques, and phage display technology.

Naïve Fab library, also known as natural Fab library, is a type of antibody library in phage display technology. The naïve libraries were constructed by cloning antibody gene fragments from natural, non immunized B lymphocytes, mainly VH-CH and VL-CL fragments, which are linked together by disulfide bonds to form Fab fragments. The naïve libraries contains a large number of antibody sequences that have not been exposed to antigens or screened by the immune system, thus possessing broad antigen binding potential and diversity.

Alpha Lifetech Can Provide

Alpha Lifetech has extensive experience in the phage display antibody discovery. Our phage display platform is well-established and we have been able to use it for various types of phages. The M13 phage platform is the most widely used for phage display. It is also extensively used in many research areas. There are a variety of phage display services available via T4 and T7 phage system, which may offer different promising options depending on their particular merits.

Alpha Lifetech has many years of experience in the field of antibodies and has built a comprehensive antibody discovery platform, ranging from monoclonal antibody discovery, polyclonal antibody development, recombinant monoclonal antibody development to various types of antibody preparation and development services. Based on different antibody preparation technologies and platforms, Alpha Lifetech provides customized services for monoclonal and polyclonal antibodies of different species. In addition to fab antibody production, we can also provide a series of fab 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. Alpha Lifetech has a variety of fab antibody purification instruments and equipment, which can provide antibody purification services from various sources such as rabbit, sheep, chicken, and mouse monoclonal antibodies, as well as Protein A/G affinity purification services and antibody separation and purification services. And we will choose the purification method based on the specific purpose of the customer. Based on the comprehensive platform system construction of antibody platform, protein platform, etc., we cover the upstream and downstream services of fab antibody production, and can provide technical services from antibody preparation, antibody purification and antibody separation and purification, antibody sequencing, antibody validation, etc.

Naïve Fab Antibody Library Construction Service

B lymphocytes are isolated from the peripheral blood of animals that have not been immunized, and because these B lymphocytes are not immunized, the antibody gene sequences they contain are diverse. The gene fragments encoding antibodies VH-CH and VL-CL were amplified from B lymphocytes by PCR, and these gene fragments contain the variable regions of the antibodies. The amplified antibody gene fragment is fused with the phage capsid protein gene, expressed, and displayed on the surface of the phage. Each bacteriophage can carry a specific antibody sequence and can be extensively amplified and screened in vitro, resulting in the antibody library construction containing a large number of different antibody sequences.

Naïve Fab Antibody Library Construction Workflow

Steps	Specification	Timeline
Step1: B Cell Extraction	Construct a diverse pool of B-cells is collected from animal blood	1 day
Step2: RNA Isolation	RNA is extracted from these B-cells, focusing on the mRNA	1 day
Step3: cDNA Synthesis	Using reverse transcription, the mRNA is converted into complementary DNA (cDNA)	1 day
Step4: Amplification of Variable Regions	The variable regions of the heavy (VH) and light (VL) chains of the antibody are then amplified using PCR	1 day
Step5: Fab Antibody Library Construction	The VH and VL regions are linked to constant regions to form the Fab fragments	2 days
Step6: Fab Antibody Library Library Cloning	Fab fragments are cloned into phage display vectors	3 days
Step7: Phage Display	The Fab library is displayed on the surface of bacteriophages, then perform high-throughput screening	1 week