Autoscanner

Single stained sections were digitally scanned using a Leica autoscanner at 20x magnification. Positive cells/mm^{2 (link)} were analyzed using ScanScope software (Leica Biosystems), or Qupath.^{23 (link)} Whole tissue regions (1 section/patient) were selected for analysis by identifying tumor cells surrounded by clear desmoplastic stroma in consultation with a pathologist (C.B.) Adjacent normal and adipose tissue on the periphery was excluded from analysis. These analysis regions also included TLS when present. Hi vs. low cutoffs were determined by median cut-points. E-TLS positivity was determined by recognition of two or more lymphoid aggregates observed to contain both CD20⁺ B cells and CD3⁺T cells. For all patients, a range of 4–8 slides were examined/tissue block that were directly serial to the original H&E used for diagnosis and the sample extracted for RNA-sequencing. These sections were used for IHC and/or additional multiplex immunofluorescence where indicated. Additionally, 5 TLS⁻ patients were randomly selected to assess for the possible presence of TLS in 3 different tissue blocks with pathologically defined cancer whereby all of the available H&E slides cut from those blocks was examined for the presence or absence of a TLS (8–13 slides/block). None of these five patients was determined to contain a TLS.