Solubilization and partial isolation of human melanoma tumor-associated antigens.
Human melanoma cell membrane tumor-associated antigens (TAA's) were solubilized in an active form by pronase digestion of either a fresh melanoma or cells from a melanoma cell line maintained in tissue culture. Upon elution from Sephadex G-200 column, TAA's solubilized from the melanoma cell line were found in four distinct peaks that had apparent molecular weights of approximately 48,000 (partition coefficient Kd, 0.426), 25,000 (Kd, 0.567)8 17,000 (Kd, 0.699), and 13,000 (Kd, 0.831) daltons, respectively. Fetal antigen activity was found in all but the 13,000-dalton peak. HLA antigen activity was detected in the 17,000-dalton material. TAA's prepared from the fresh tumor source eluted from Sephadex G-200 column with an apparent molecular weight of 14,000-25,000 (Kd, 0.786-0.572) daltons, as did HLA antigens. A partial resolution of the TAA's from the HLA antigens was achieved with the use of DEAE-cellulose chromatography. Results of antigenic stability assays suggested that the TAA structure is stable to prolonged exposure to low pH. Recovery of TAA activity from the strong denaturing agents 5 m urea, 0.5% (wt/vol) sodium dodecyl sulfate, and 4 m guanidine hydrochloride was partially successful. These properties of the TAA's may be useful for further isolation of the TAA's.
Stuhlmiller, GM; Green, RW; Seigler, HF
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