Objectives: This study aimed to examine (1) the expression of P16 protein relative to sites of presentation, immunophenotypic subgroups and proliferative indices of tumour cells, and (2) the relationship between p16 gene alterations and P16 protein overexpression in 70 cases of diffuse large B cell lymphoma (DLBCL). Methods: Expression of P16, CD10, BCL-6, MUM-1 and proliferation marker (Ki-67) was demonstrated by immunohistochemistry. Fluorescence in situ hybridization (FISH) was employed to detect p16 alterations. Results: P16 overexpression was shown in 45.7% (32/70) of the DLBCL cases, and was significantly correlated with CD10 (p = 0.022) and germinal centre B-cell-like (GCB) phenotype (p = 0.022). High expression of P16 was inversely associated with high proliferative activity (Ki-67 index greater than 75%) (p = 0.020). Of the 47 cases that yielded interpretable FISH results, 57.4% (27/47) showed deletions of p16 and 27.7% (13/47) showed gains of p16. P16 overexpression and p16 deletions were mutually exclusive (p = 0.019). There was no correlation between P16 overexpression and p16 gains (p = 0.621). Conclusions: The GCB and non-GCB subgroups of DLBCLs show different patterns of P16 expression. High levels of P16 may mitigate tumour cell proliferation. Gains of p16 do not necessarily increase P16 protein expression.