The diagnostic value of the generally accepted Limulus lysate endotoxin test was examined in a series of pediatric cancer patients having or suspected of having Gram-negative bacterial sepsis. One hundred fifty-four tests were performed, with simultaneous blood cultures available for 143 of these. Of the 28 patients with one or more separate episodes of Gram-negative bacillary bacteremia, only two had positive endotoxin tests. Eight children who died with Gram-negative septicemia within 24 hours after being tested for endotoxin had negative tests. These results, supported by others, indicate that the Limulus amebocyte lysate test is not clinically useful for detecting Gram-negative infections or predicting their outcome. Lack of standardized procedures for lysate preparation and endotoxin extraction from plasma could explain the current disagreement of published test results.