Optimising protein detection with fixable custom oligo-labelled antibodies for single-cell multi-omics approaches

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http://hdl.handle.net/10138/346129

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Kleino , I , Nowlan , K , Kotimaa , J & Kekäläinen , E 2022 , ' Optimising protein detection with fixable custom oligo-labelled antibodies for single-cell multi-omics approaches ' , Biotechnology Journal , vol. 17 , no. 6 , 2100213 . https://doi.org/10.1002/biot.202100213

Title: Optimising protein detection with fixable custom oligo-labelled antibodies for single-cell multi-omics approaches
Author: Kleino, Iivari; Nowlan, Kirsten; Kotimaa, Juha; Kekäläinen, Eliisa
Contributor organization: TRIMM - Translational Immunology Research Program
Medicum
Department of Bacteriology and Immunology
HUSLAB
Date: 2022-06
Language: eng
Number of pages: 11
Belongs to series: Biotechnology Journal
ISSN: 1860-6768
DOI: https://doi.org/10.1002/biot.202100213
URI: http://hdl.handle.net/10138/346129
Abstract: Background and Aim Single-cell RNA sequencing (scRNA-seq) is a powerful method utilising transcriptomic data for detailed characterisation of heterogeneous cell populations. The use of oligonucleotide-labelled antibodies for targeted proteomics addresses the shortcomings of the scRNA-seq-only based approach by improving detection of low expressing targets. However, optimisation of large antibody panels is challenging and depends on the availability of co-functioning oligonucleotide-labelled antibodies. Main Methods and Results We present here a simple adjustable oligonucleotide-antibody conjugation method which enables a desired level of oligo-conjugation per antibody. The mean labelling in the produced antibody batches varied from 1 to 6 oligos per antibody. In the scRNA-seq multimodal experiment, the highest sensitivity was seen with moderate antibody labelling as the high activation and/or labelling was detrimental to antibody performance. The conjugates were also tested for compatibility with the fixation and freeze storage protocols. The oligo-antibody signal was stable in fixed cells indicating the feasibility of a stain, fix, store, and analyse later type of workflow for multimodal scRNA-seq. Conclusions and Implications Optimised oligo-labelling will improve detection of weak protein targets in scRNA-seq multimodal experiments and reduce sequencing costs due to a more balanced amplification of different antibody signals in CITE-seq libraries. Furthermore, the use of a pre-stain, fix, run later protocol will allow for flexibility, facilitate sample pooling, and ease logistics in scRNA-seq multimodal experiments.
Subject: antibody conjugation
CITE-seq
single-cell RNA-seq
CD38
ACTIVATION
3111 Biomedicine
1182 Biochemistry, cell and molecular biology
Peer reviewed: Yes
Rights: cc_by_nc_nd
Usage restriction: openAccess
Self-archived version: publishedVersion


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