Primary human squamous-cell lung tumours were disrupted using mechanical high-speed homogenization. Crude membranes were isolated by differential centrifugation at low speeds followed by 100, 000 g centrifugation. Such membrane preparations were extracted with Triton-X-100 and passed over a DEAE cellulose column, the DEAE unbound fraction and the bound fraction eluted with 0·4M NaCl were used to immunize rabbits. We present here only data on a single lung-tumour-associated membrane antigen (TAMA) found in the unbound DEAE cellulose column eluates. This new Triton-X-100 extractable antigen is termed lung TAMA-1.A further purification of the antigen was achieved using two methods. The first used G-200 molecular-seive chromatography and this revealed lung TAMA-1 to have a mol. wt of 200, 000. A second method used sucrose density-gradient isoelectric focusing, and the antigen had an isoelectric point of ∼3·0.Several important properties of the lung TAMA-1 were determined. The antigen has a cathodal gamma-type electrophoretic mobility at pH 7·6. The antigen was not detected in any normal human adult or foetal tissue extracts tested. It did not crossreact immunochemically with CEA, AFP or β2 microglobulin. Lung TAMA-1 was detected in 80% of lung tumour Triton-X-100 extracts tested by counterimmuno-electrophoresis (CIEP) but was undetectable in breast or colon carcinoma extracts. Low frequency sonication did not deleteriously affect lung TAMA-1, but, 3M KCl eliminated its immunologic reactivity in CIEP. Finally, preliminary data were obtained using immunohistochemistry to localize in vivo lung TAMA-1 production.
ASJC Scopus subject areas
- Cancer Research