We occasionally are asked about the precise meaning of all the fields in customer data files (.tsv) downloaded from the immunoSEQ website. Perhaps the easiest way to explain this is a table with the name of each field along with a definition.
| Column Title | Definition |
| sequenceID | This is the flow cell lane_sample ID_primer versions |
| container | Project ID associated with the samples |
| nucleotide | Reconstructed nucleotide sequence |
| aminoAcid | Inferred amino acid sequence |
| normalizedFrequency | Frequency calculated based on normalized copy number *PCR normalization is currently considered reliable only for Human TCR beta |
| normalizedCopy | Each copy count is multiplied by a factor depending on the V-J pair identity, to account for V and J primer bias. *For Human TCR beta |
| rawFrequency | Frequency calculated based on raw copy number |
| copy | The raw copy count |
| cdr3Length | Length of the CDR3 region-including indels |
| VFamilyName | V family number if known (i.e. including if there are ties) |
| VGeneName | V family number and specific V gene number-or if known to either of two related genes-or unknown if more |
| VTies | Names of potential V genes |
| DGeneName | D gene name including unknowns |
| JGeneName | Name of J gene. If a J tie, returns the alphabetically first J gene name |
| JTies | Names of potential J genes |
| VDeletion | The number of deletions inferred after V identification |
| d5Deletion | The number of deletions between the end of the V segment and the 5′ end of the D segment |
| d3Deletion | The number of deletions between the end of the J segment and the 3′ end of the D segment |
| JDeletion | The number of deletions inferred after J identification |
| n2Insertion | The number of insertions in the N2 region |
| n1Insertion | The number of insertions in the N1 region |
| sequenceStatus | Whether the nucleotide sequence generates a functional amino acid sequence |
| VIndex | Distance from the start of the V gene (designated 0) to a conserved V motif |
| n1Index | Distance from 0 to the start of the N1 region |
| n2Index | Distance from 0 to the start of the N2 region |
| DIndex | Distance from 0 to the start of the D region |
| JIndex | Distance from 0 to the 3′ end of the J region |
Below is a cartoon illustrating the V,D,J, and N index positions.
As for the difference between “raw” and “normalized,” the set of normalization factors we currently feature with our online tools is derived from comparing sequence data to results from a Betamark (flow) assay that measures relative amounts of each V. The results for most V’s matched quite closely but any remaining differences are fixed by the multiplication of V-J specific normalization factors.
