The Application of Single-Cell RNA Sequencing in Vaccinology

Box 2: scRNA-seq as a starting point for monoclonal antibody production.

Recently, efforts to produce and characterise monoclonal antibodies (mAb) have made impressive progress. mAbs with broadly neutralising activity against specific antigens largely act through their Fab fragment specificities.

Our comprehension of the way antibody specificities interact with B cell function has remained limited due to the intricacies of polyclonal antibody responses. Neu et al. developed the Spec-seq protocol to tackle this challenge ([92]; Table 1). Spec-seq permits mAb production and transcriptional profiling from the same cell. This involves modifications to the Smart-seq2 scRNA-seq method [93] and the harmonisation of two mAb generation techniques [94, 95]. The authors apply Spec-seq to analyse plasmablasts following influenza vaccination, characterising differences in transcriptome according to BCR isotype and vaccine reactivity. IgA vaccine-reactive circulating plasmablasts were compared with IgG vaccine-reactive and IgA vaccine-nonreactive populations. The authors suggest that imprinting following initial B cell activation results in transcriptional similarities among clonally related plasmablasts. Total and IgA plasmablasts cluster in tSNE projections by their ability to bind the vaccine, suggesting transcriptional modules correlated with vaccine reactivity. These data highlight an unexpectedly high degree of transcriptional specialisation within plasmablasts, demonstrating that these cells are not terminally differentiated antibody-secreting cells. Spec-seq is a robust technique that allows the profiling of the transcriptome and mAbs of hundreds of cells.

With improvements in droplet-based scRNA-seq methods, the scale (thousands of single cells) and order of events (BCR sequence and transcriptome information first, mAb generation thereafter) could be changed. Rather than characterising every mAb “blind” (i.e., without any prior information on its cell of origin), mAbs could be selected on the basis of transcriptome information and clonal family position in a hope to only generate and characterise high affinity/avidity candidates. Combining this with fluorescence-assisted cell sorting (FACS) to isolate vaccine antigen-specific B cells could provide a powerful new workflow to produce monoclonal antibodies against specific pathogens. An example of this type of workflow is put forward by Goldstein and colleagues [96] and is already being put to use for mAb development in priority diseases (see Box 3) [97].

Box 2: scRNA-seq as a starting point for monoclonal antibody production.