Construction Of Na?ve And Immune Nanobody Libraries Using Alpaca (Lama Pacos) As A Convenient Source

Antibodies are the antigen-binding immunoglobulins secreted by plasma cells.Nanobodies,also called recombinant single-domain antibody fragments,are the smallest naturally integral antigen-binding units.Nanobodies are derived from the heavy-chain antibodies of camelid.Compared with common antibodies,they have smaller molecular weight,higher affinity,better tissue penetration,lower immunogenicity,and can be genetically manipulated more easily.These properties make them robust tools for biomedical applications,and might take the place of conventional antibodies in the near future.Phage display is the most wildly used method to obtain nanobodies.With this technique,the gene encoding target protein is merged with the phage capsid protein gene,making the protein displayed on the surface of phage.After the process of biopanning,our target nanobodies can be screened and amplified for economic benefit.Classical antibodies are derived from peripheral blood mononuclear cells of human or mouse,while nanobodies are derived from peripheral blood lymphocytes of camels and llamas.Alpaca,a kind of raising friendly camelid animal,is considered to be the best source of nanobody.However,while nanobodies derived from alpaca have become widely used in research beyond seas,they are not well studied in China.Here,we constructed large and diverse phage display nanobody libraries from na?ve(with library size of 1.4*109 c.f.u.)and GFP-Flag-His immunized(with library size of 1.7*1010 c.f.u.)alpaca.Theoretically,these two nanobody libraries can be used for the screening of any nanobodies.Meanwhile,we have finished the preliminary screening using GFP-Flag-His immune library,and obtained two protein sequence enrichment.The results demonstrates that this library can indeed be used to produce nanobodies,and phage display platform has been successfully established.At present,commercial antibodies against human telomere binding proteins are short of affinity or specificity.Up to now,we have purified enough amount of human telomere binding proteins,including TRF1,TRF2 and RAP1.Then specific human telomere binding protein nanobodies can be selected and amplified.These nanobodies can be modified for specific application and obtain higher titers and more economic benefits than conventional antibodies.For the study of telomere,it may become a superexcellent research tool in years to come.