Abstract
BackgroundRepertoire analysis of patient-derived recombinant monoclonal antibodies is an important tool to study the role of B cells in autoimmune diseases of the human brain and beyond. Current protocols for generation of patient-derived recombinant monoclonal antibody libraries are time-consuming and contain repetitive steps, some of which can be assisted with the help of software automation.ResultsWe developed BASE, an easy-to-use software for complete data analysis in single cell immunoglobulin cloning. BASE consists of two modules: aBASE for immunological annotations and cloning primer lookup, and cBASE for plasmid sequence identity confirmation before expression. Comparing automated BASE analysis with manual analysis we confirmed the validity of BASE output: identity between manual and automated aBASE analysis was 100% for all outputs, except for immunoglobulin isotype determination. In this case, aBASE yielded correct results in 96% of cases, whereas 4% of cases required manual confirmation. cBASE automatically concluded expression recommendations in 89.8% of cases, 91.8% of which were identical to manually derived results and none of them were false-positive.ConclusionsBASE offers an easy-to-use software solution suitable for complete Ig sequence data analysis and tracking during recombinant mAb cloning from single cells. Plasmid sequence identity confirmation by cBASE offers functionality not provided by existing software solutions in the field and will help to reduce time-consuming steps of the monoclonal antibody generation workflow. BASE can be installed locally or accessed online at Code Ocean.
Highlights
Repertoire analysis of patient-derived recombinant monoclonal antibodies is an important tool to study the role of B cells in autoimmune diseases of the human brain and beyond
The data set is derived from a cerebrospinal fluid (CSF) cell sample processed using monoclonal antibodies (mAb) repertoire cloning in our laboratory (sample ID #AI ENC 113, (Kreye J, et al.: Encephalitis patient derived monoclonal GABAA receptor antibodies cause catatonia and epileptic seizures, in preparation)) including a total of 181 amplified cDNA Ig sequence reads for aBASE validation and a total of 176 plasmid Ig sequence reads for cBASE validation
Validation of primary Ig sequence evaluation using aBASE Comparison of the results of the analysis performed by aBASE and the manual analysis expectedly showed no relevant differences in the parameters belonging to preprocessing analysis and the data extracted from IgBLAST (Table 1)
Summary
Repertoire analysis of patient-derived recombinant monoclonal antibodies is an important tool to study the role of B cells in autoimmune diseases of the human brain and beyond. Repertoire analysis of patient-derived recombinant monoclonal antibodies (mAb) has become a state-of-the-art approach to investigate B cell (patho-)physiology and assess the role of (auto-)antibodies in disease progression in a variety of autoimmune and infectious diseases [1,2,3,4,5,6,7]. Recombinant expression approaches can generate libraries of purified mAbs which can be directly used in functional assays and further downstream applications. In neuroscience, applying these approaches to cerebrospinal fluid (CSF)-derived cells has opened a rapidly developing field: Studying central nervous system mAb repertoires (brain antibody-omics) enables insights into the physiological or pathogenic role of (auto-)antibodies in health and disease. Protocols require optimization to increase successful expression rates from derived single cells in comparison to bulk approaches from blood samples
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
