Lc817

Approval to use human specimens was given by the Institutional Review Board of Emory University. All clinical samples were collected with informed consent under Health Insurance Portability and Accountability Act (HIPAA) approved protocols. Paraffin-embedded lung cancer patient tumors were obtained from US Biomax (LC814 and LC817).

Paraffin-embedded lung cancer tissue microarrays (LC814 and LC817) containing primary and matched metastasized tumors from lymph nodes were obtained from US Biomax. IHC analyses were performed according to the previously described (Kang et al., 2010 (link)). In brief, human tissue sections were deparaffinized, rehydrated, and incubated in 3% hydrogen peroxide to suppress endogenous peroxidase activity. Antigen retrieval was achieved by microwaving the sections in 10 mM Sodium Citrate (pH 6.0). Sections were incubation in 2.5% horse serum for blocking. The primary antibodies were applied to the slides at dilution of 1:250 (anti-LKB1 antibody), 1:200 (anti-PLAG1 antibody), 1:500 (anti-GDH1 antibody), and 1:100 (anti-p-AMPK T172 antibody) at 4°C overnight. Detection was achieved with the avidin–biotin complex system (Vector Laboratories). Slides were stained with 3,3′-diaminobenzidine, washed, counterstained with hematoxylin, dehydrated, treated with xylene, and mounted. Positive staining was identified using IHC signal intensity scored as 0, +1, +2, and +3.