The addition of gelatin to blood culture media has been suggested to prevent the inhibition of Neisseria meningitidis, Neisseria gonorrhoeae, Gardnerella vaginalis, and Peptostreptococcus anaerobius that is caused by sodium polyanetholsulfonate. To determine the effect of such supplementation on the overall yield of microorganisms, we compared the yield and speed of detection of clinically important microorganisms from 5422 paired 10-ml samples of blood cultured in Trypticase soy broth (TSB) containing 0.03% sodium polyanetholesulfonate (SPS) and TSB/SPS containing 1.2% gelatin and 1.0% yeast extract (mTSB). The atmosphere of incubation (open venting unit) and ratio of blood to broth (1:5) were the same for both samples. Only cultures with adequate blood sample (greater than or equal to 80% of stated volume) were compared statistically. Addition of gelatin and yeast extract resulted in inhibited growth of Enterobacteriaceae (p less than 0.001), Pseudomonas aeruginosa (p less than 0.01), fungi (p less than 0.05), and the overall set of microorganisms encountered (p less than 0.001). It delayed growth of Enterobacteriaceae (p less than 0.001) but reduced the time to recover staphylococci (p less than 0.02). Of 12 isolates of species usually inhibited by SPS, seven grew only with the addition of gelatin and yeast extract, none grew only without supplementation, and five grew in both media. Although gelatin and yeast extract may improve the yield of some specific bacteria, the routine use of these additives cannot be recommended for all blood culture media.