Proprietary phagemid vector
WizAmpTM single-primer method amplifies antibody genes uniformly
We have developed a patented antibody library construction method termed WizAmpTM (US 9,890,414), which enables more uniform and robust amplification of antibody genes compared to traditional 2-primer PCR methods. In WizAmpTM, mRNA is purified from homogenized tissue and first-strand cDNA is first synthesized using a proprietary primer mixture recognizing rabbit antibody genes (see below). During second strand cDNA synthesis, matching nucleotide sequences at both 5’ and 3’ ends are appended, allowing for subsequent antibody gene amplification using a single non-gene-specific primer.
Traditional library construction methods employ 2-primer PCR. The use of two primers with mismatching sequences and even slight differences in melting temperature can cause non-specific and uneven amplification that will degrade the quality of the library. Unbiased, robust amplification by a single single non-gene-specific primer creates high quality libraries that preserve that natural frequency and proportion of each antibody gene isolated from the immunized rabbit.
WizAmpTM recovers maximal antibody gene diversity from immune sources
Through iterative improvement after creating hundreds of rabbit libraries, we have created a primer set that covers the highest reported rabbit VH FR1 sequence diversity to date by including many FR1 sequences that are not currently in publicly available databases. These primers initiate first strand cDNA synthesis using mRNA purified from homogenized immune tissue, prior to the single-primer amplification step above. Compared to literature sources or a competitors’ published primer set, our primers cover many more rabbit antibody genes.
This primer set was developed iteratively – we have observed for a variety of antigens that when only a partial set of 32 primers are used for library construction, no target-specific antibodies can be isolated after phage panning. However, when the full set of 39 primers was used for library construction from the exact same rabbit spleen source, dozens of hits were obtained. Each immune response is unique – without full coverage of the immune repertoire, a target-specific antibody may be missed.
See how we pair these technologies with our proprietary phagemid vector to deliver rabbit Fab phage display for the first time.
See how we have used this cutting-edge library construction technology to build rabbit immune libraries to isolate rare, functional mAbs against GPCRs and ion channels and to make highly-specific compantion diagnostic rabbit monoclonal antibodies.