Measurement of endotoxin activity in critically ill patients using whole blood neutrophil dependent chemiluminescence

Abstract

Background: Lipopolysaccharide (endotoxin) from the cell wall of Gram-negative bacteria is a potent trigger for the release of host-derived inflammatory mediators. The relationship between endotoxaemia, Gram-negative infection and the clinical syndrome of sepsis has been difficult to establish, in part because of the limitations of available endotoxin assays.

Methods: We performed an observational cohort study in critically ill patients in the medical/surgical intensive care unit (ICU) of a tertiary care hospital. Whole blood endotoxin levels on the day of ICU admission were measured using a novel chemiluminescent assay--the endotoxin activity assay (EAA)--and the chromogenic modification of the limulus amoebocyte lysate (LAL) assay.

Results: We studied 74 consecutive admissions. Endotoxin levels were higher in patients with a diagnosis of sepsis (470 +/- 57 pg/ml) than in patients admitted with a diagnosis other than sepsis (157 +/- 140 pg/ml; P < 0.001). Endotoxaemia was significantly associated with Gram-negative infection (P < 0.05); no patient with a Gram-negative infection had an endotoxin level below 50 pg/ml. White blood cell counts of patients with EAA-detected endotoxaemia were significantly higher (15.7 +/- 9.1 x 10(9) cells/l for endotoxaemic patients versus 10.8 +/- 6.2 x 10(9) cells/l for patients without endotoxaemia; P < 0.05).

Conclusion: Endotoxaemia is associated with Gram-negative infection from any source, and with a diagnosis of sepsis and leukocytosis. These correlations were not apparent using the LAL method. The EAA may be a useful diagnostic tool for the investigation of invasive Gram-negative infection and incipient sepsis.

Figure 1

Effect of polymyxin-sulphate preincubation on endotoxin recovery in the endotoxin activity assay. Samples from 15 intensive care unit patients with endotoxaemia ranging from 50 to more than 800 pg/ml were pretreated with polymyxin-sulphate at a concentration of 300 g/ml for 20 min at 37°C. Endotoxin concentrations are expressed as means ± standard deviation.

Figure 2

Endotoxin levels in patients with Gram-negative infection. Endotoxin levels were significantly higher by endotoxin activity assay (P < 0.05, n = 10). Endotoxin concentrations are expressed as means ± standard deviation.