Screening And Recombinant Expression Of Anti-HSA Monoclonal Antibody

Human serum albumin(HSA)is the most abundant protein in the plasma and is mainly produced by the liver.It plays an important role in maintaining the normal function of the body and treating related diseases.At the same time,HSA is also an important biological indicator for many diseases.For the methods of HSA detection,there are mainly radioimmunoassay,enzyme-linked immunosorbent assay,immunoturbidimetry,etc.These methods often rely on the specific binding of antibody and antigen to achieve accuracy,specificity and sensitivity of HSA detection.Especially when used in complex environments or trace detection,the antibodies used must have stable and reliable properties.Therefore,it is of great significance to obtain anti-HSA monoclonal antibodies(mAb)with high specificity and binding ability.And,analyzing or modifying the full-length gene of anti-HSA mAb helps HSA detection and diagnosis of related diseases.The acquisition of antibody variable regions or full-length information is essential for antibody engineering work and antibody function identification.This study defined a workflow,which allows researchers to identify the full-length immunoglobulin(Ig)gene sequence of monoclonal antibodies with high accuracy from a single hybridoma cell.The main contents of this study:The heavy/light chain(HC/LC)gene sequences of the mAb against HSA were identified by using Smart-seq2 followed by BASIC?rnaSPAdes and Igfinder.The antibody variable regions were cloned using traditional RT-PCR technology to verify the results of single-cell transcriptome analysis.The results show that the Ig gene sequence obtained by single-cell transcriptome analysis is identical to the sequence obtained by RT-PCR.After that,the obtained Ig genes were cloned into eukaryotic expression plasmids and expressed in HEK293 T cells.The results show that purified recombinant antibody specifically recognizes HSA and has strong binding ability to it.All this proves that the monoclonal antibody sequence obtained by our method is reliable.In summary,this study describes a method with high-accuracy that allow identification the HC and LC gene sequences of anti-HSA mAb from a single hybridoma cell.We expect that this method can also be applied to other hybridoma cell lines or single lymphocytes isolated from other species,which will help to identify the characteristics of antibodies and promote the development of antibody engineering.