Use of avidin to prepare biotin-free culture media

Dialysis of avidin-treated casein hydrolysate and peptone solutions produced biotin-free dialysates which, when incorporated into growth media resulted in media devoid of biotin. Wright-Skeggs medium supplemented with dialyzed, avidin-treated, casein hydrolysate did not support significant growth of Bacillus coagulans or Lactobacillus plantarum without biotin supplementation. Eagle's minimal essential medium supplemented with avidin-treated, peptone dialysate (MEMAPD) would not support the growth of LM strain mouse cells through more than a single passage without biotin supplementation. MEMAPD supplemented with biotin supported good growth of the LM cells through repeated passages. Using this medium it was possible to demonstrate that LM cells require biotin for growth and that they cannot utilize avidin-complexed biotin. Avidin treatment of peptone or casein hydrolysate solutions allowed accurate control of the biotin content in culture media and did not remove significant amounts of substances other than biotin. By comparison, activated charcoal nonspecifically removes biotin as well as other essential nutrients and adds unknown substances to the medium which interfere with the growth of tissue culture cells.