Question: What is the limit of detection for a specific clonotype using the Single Cell Immune Profiling assay?
Answer: In our internal testing, we have estimated that the limit of detection for a specific clonotype is approximately 1%, although this will depend on the expression level of the V(D)J transcripts within the cell.
To test this, we created a cell mixture of standard PBMCs with a 1% spike in of EBV-specific T cells. We targeted both 10,000 total cells and 5,000 total cells. From this, we obtained ~4000 T cells and ~2000 T cells, respectively. We then investigated the T cell data to see if we could observe the EBV-specific clonotypes we had spiked in.
Of the EBV-specific cells, we knew that around ~55% expressed the following major clonotype: TRAV12-3: J20 (CDR3: CATQGSNDYKLSF), TRBV9:D1: J1-4 (CDR3: CASSTGQVATNEKLFF), so we would expect to be able to detect this clonotype in the data at approximately 0.55% of the population (1% of 55%).
We were able to detect this major clonotype in the cell suspension, with fewer than 10 cell expressing it in the population.
There was also a minor clonotype in the EBV-specific population that was expressed at ~9% of the population: TRAV5: J15 (CDR3: CAESSNQAGTALIF), TRBV14:D2: J2-1 (CDR3: CASSQSPGGIQFF), so we would expect to be able to detect this clonotype in the data at approximately 0.09% of the population (1% of 9%).
We were also able to detect this minor clonotype at approximately the expected levels in both experiments, although in the 5000 cell experiment, only the beta chain was detected.
Products: Single Cell Immune Profiling