Product Overview
Fusion BioLabs offers a range of library primer sets and phagemid vector combinations for antibody phage display and peptide phage display construction. With customizable features and robust performance, our primer sets and phagemid vectors are designed for facilitating phage display library generation as fast as within one week.
pAPD-m-Fab is the phagemid vector for construction of a fragment antigen-binding (Fab) library for mouse antibodies. Here are the key steps involved in constructing such a library:
- Amplify V genes from cDNA reverse transcript from RNA isolated from peripheral blood lymphocytes (PBL) or lymphoid tissue of non-immunized or immunized donors using PCR primers corresponding to known VH, Vκ, and Vλ gene sequences.
- Combine VH repertoires and the CH1 fragment and VL repertoires and the CL fragment to create VH-CH1 and Vk,λ-Ck,λ constructs, respectively, using a simple two-fragment PCR assembly procedure.
- Restriction enzyme digestion of the pAPD-m-Fab vector and Vk,λ-Ck,λ fragments with SacI/XbaI, or pAPD-m-Fab vector and VH-CH1 fragments with XhoI/SpeI.
- Ligation of the digested and purified fragment into the corresponding restriction enzyme-digested and purified pAPD-m-Fab vector to make either a light chain sub-library or a heavy chain sub-library.
- Restriction enzyme digestion of the light chain sub-library and VH-CH1 fragment with XhoI/SpeI or the heavy chain sub-library and Vk,λ-Ck,λ fragment with SacI/XbaI to make mouse Fab libraries.
Key Features
High expression efficiency: Engineered for efficient expression and display of antibody fragment scFv on the surface, allowing for easy screening and selection of target molecules.
Flexibility and versatility: One vector for both antibody library construction and downstream antibody fragment expression. No need to subclone into an expression vector for downstream application.
Specifications of Antibody Phage Display Vector
| Antibiotic Resistance | Ampicillin (AmpR) |
| Constitutive or Inducible System | Inducible for downstream expression |
| Delivery Type | Transformation |
| Product Type | Phage display vector or Bacterial Expression vector |
| Cloning Method | Either [double 5’end and 3’end SfiI sites] or [5’end SacI and 3’ end SpeI sites] |
Contents & Storage
Content of mouse Fab phage display library construction Kit| Primer Set | ||
| Vial 1 | 100 µl, 10 µM | Primer mix (F+R) for Vk repertoire amplification |
| Vial 2 | 100 µl, 10 µM | Primer mix (F+R) for Vλ repertoire amplification |
| Vial 3 | 100 µl, 10 µM | Primer mix (F+R) for VH repertoire amplification |
| Vial 4 | 100 µl, 10 µM | Primer mix (F+R) for Ck fragment amplification |
| Vial 5 | 100 µl, 10 µM | Primer mix (F+R) for Cλ fragment amplification |
| Vial 6 | 100 µl, 10 µM | Primer mix (F+R) for CH1 fragment amplification |
| Vial 7 | 100 µl, 10 µM | Assembly Primer mix (F+R) for Vk-Ck repertoire construction |
| Vial 8 | 100 µl, 10 µM | Assembly Primer mix (F+R) for Vλ-Cλ repertoire construction |
| Vial 9 | 100 µl, 10 µM | Assembly Primer mix (F+R) for VH-CCH1 repertoire construction |
| Sequencing Primer set | ||
| Vial 10 | 100 µl, 10 µM | M13 Reverse primer for scFv forward sequencing |
| Vial 11 | 100 µl, 10 µM | pIII Reverse primer for scFv reverse sequencing |
| pAPD-m-Fab vector for mouse Fab phage display library construction | ||
| Vial 12 | 10.0 µg in Tris-EDTA buffer | |
- Store at -20°. Vectors are guaranteed stable for 12 months when properly stored.
Vector usage
- Cloning Fab inserts into pAPD-m-Fab vector for phage display library construction.
- Cloning the Fab candidate into pAPD-m-Fab vector for antibody fragment expression.
