Arid1a

Immunohistochemistry was performed on the DAKO Autostainer (DAKO, Carpinteria, CA) using DAKO Envision+ and diaminobenzadine as the chromogen. Sections of de-paraffinized Orbital and ocular adnexal lymphomas were labeled with ARID1A (mouse monoclonal, clone PSG3, 1:250, Santa Cruz Biotechnology, Dallas TX, #sc-32761), EZH2 (mouse monoclonal, clone 11, 1:100, BD Biosciences, San Jose, CA, #612666), or KMT3B/NSD1 (rabbit polyclonal, 1:250, EMD Millipore, Billerica, MA, #ABE1009) for 60 minutes at ambient temperature. Microwave epitope retrieval in 10 mM Tris/HCl, pH 9 containing 1 mM EDTA was used prior to staining. Appropriate negative (no primary antibody) and positive controls were stained in parallel with each set of tumors studied. Immunostained slides were examined by light microscopy by two pathologists (A.S.M. and S.A.T.). Only staining of the nucleus was marked as positive expression. The staining was scored semiquantitatively and recorded based on percent nuclei staining (0 = negative, 1 = 1 – 25% immunoreactive cells, 2 = 26% – 50% immunoreactive cells, 3 = 51% – 75% immunoreactive cells, 4 = 76% – 100% immunoreactive cells) and intensity of staining (0 = negative, 1 = weak, 2 = moderate, 3 = strong). Corresponding sections were also H&E stained.

ChIP was performed as we have previously described ^{19 (link)}. The following antibodies were used to perform ChIP: ARID1A (Santa Cruz), RNA polymerase II (Santa Cruz), H3Ac (Active Motif) and BRG1 (Santa Cruz). An isotype matched IgG was used as a negative control. ChIP DNA was analysed by quantitative PCR against the promoter of the human HDAC6 gene. All primer sequences in Table S1. For single site PCR, the primers for position −880 upstream of transcription starting site were used.