Set A (128 tumors): For comparison of RPPA with transcriptional profiling (e.g., for protein–mRNA correlations), 128 stored primary breast tumors were obtained from patients treated in the Danish DBCG82 b and c studies [45 (link)] (
Set B (ten tumors): For the studies of intratumoral heterogeneity and total and phosphoprotein stability, a prospective study was undertaken to collect primary breast tissue at breast surgery in ten patients with breast cancer under an Institutional Review Board (IRB)-approved protocol. Each tumor was sectioned with assistance from a breast pathologist and immediately snap frozen (three pieces) or left at room temperature in closed eppendorf tubes without any added buffer for 0.5/1/2/4/6/24 h (1 piece/time point) prior to freezing (−85°C). Protein was extracted from each piece of tumor without thawing.
Set C (95 tumors): Ninety-five stored primary breast tumors were obtained from the breast tumor frozen tissue bank at M. D. Anderson Cancer Center under an IRB-approved protocol (
MDAMB231 and MDAMB435 breast cancer xenografts were assessed for total and phosphoprotein stability using the same approach as with human tumor set B above. After animal sacrifice, the xenograft tumors were sectioned and immediately snap frozen or left at room temperature in closed eppendorf tubes without any added buffer for 0.5/1/ 2/4/6 h (1 piece/time point) prior to freezing (−85°C). As with the human tumors, protein was extracted from each piece of tumor without thawing.