The amount and nature of lipids that accumulate in "old" cultures of chick embryo heart fibroblasts were studied, along with the effect of vitamin B12 and folic acid on the accumulation. A technique was presented for separation of outgrowth tissue from tissue implanted for analysis. Vitamin B12 and folic acid in the medium prevented accumulation of fat and apparently stimulated growth of the culture. Normal controls, freshly excised tissue, and vitamin-treated cultures were essentially free of neutral fat, as indicated by constancy of the molar ratio of fatty acids to phospholipid P, approximately 2. In degenerating cultures this ratio increased rapidly in the absence of added vitamins.