Title:

Anti-HIV Antibody Optimization via Site Saturation Mutagenesis

Human immunodeficiency virus (HIV) causes over one million annual deaths globally and lacks an effective vaccine. Antiretroviral therapy (ART) is the predominant form of treatment, but it is often inaccessible due to limited healthcare infrastructure and cost in many developing nations. Monoclonal antibody (mAb) cocktails present a viable treatment alternative that is urgently needed. VRC07-523LS-FR303 is a a broadly neutralizing mAb (bNAb) developed at the NIH with high neutralization breadth and potency against a 208-virus panel. To further improve breadth and potency, substitution, insertion, and deletion oligonucleotide primers were designed and used to perform NNK and MNN site saturation mutagenesis. PCR annealing temperatures were adjusted to achieve optimal mutation efficiencies. The resultant VRC07-523LS-FR303 mutant plasmids will be transformed into yeast display libraries for future fluorescence-activated cell sorting and single-cell analysis.

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