Measuring Endotoxin:

Spectral’s EAA™ is a novel method to measure endotoxin. Numerous previous attempts to develop a reliable and accurate assay for endotoxin have been unsuccessful, mainly due to the complexity of the interaction of endotoxin with other elements in the blood such as binding proteins. Scientists at Spectral have incorporated the body’s natural mechanisms of endotoxin detection and adapted them into a system that provides an accurate endotoxin measurement within 30 minutes.

The EAA™ is a chemiluminescent bio-assay based on the oxidative burst reaction of activated neutrophils to complement coated LPS-IgM immune complexes. The IgM antibody, a key reagent, is specific for the Lipid A portion of endotoxin (LPS). In the presence of LPS, the ensuing oxidative burst results in light emission in the presence of luminol. Each EAA test relies on 3 reactions: the first reaction is a negative control which allows each patient to be their own control, the second is the test sample and the third is a maximum chemiluminescence calibrator with a high level of exogenous endotoxin. The maximum calibrator is necessary as endotoxins from different species of Gram negative bacteria have a slightly different reactivity in the EAA. E. coli O55:B5 is used as the standard as it has a mid-level reactivity in the assay.

Measuring Endotoxin

Removing Endotoxin: Removing Endotoxin

Among the various strategies studied to eliminate endotoxin, Polymyxin-B hemoperfusion has been the most effective in blocking the toxic effects of endotoxin by virtue of its electrochemical affinity with Lipid A (common to all species of endotoxin). However, systemic use of Polymyxin-B is limited by severe nephro- and neurotoxicity.

The interaction between Polymyxin-B and Lipid A of endotoxin is governed primarily by forces which include:

Ionic: guides the formation of the link/bond;

Hydrophobic: breaks the spatial order of the acyl chains of LPS and neutralizes the toxicity, transforming LPS and Polymyxin-B into a single unimolecular compound.

PMX (Toraymyxin hemoperfusion) removes endotoxin from the blood using the electrochemical properties of Polymyxin-B, while avoiding known side effects of its systemic administration. The PMX cartridge contains polystyrene composite woven fibre with Polymyxin-B immobilized on the surface through covalent bonding. The blood flows radially and uniformly through the fibres at a rate of 80-120 ml/min before exiting the cartridge. The recommended dosing of hemoperfusion is two 2-hour hemoperfusion events in a 24 hour period. The in-vitro adsorption capacity (human blood) was measured as 20 μg (Romaschin et al., 2017).

PMX selectively adsorbs circulating endotoxin, thereby helping to rebalance the innate immune system. Removal of endotoxin leads to a decrease in inflammatory mediator levels (shown in the figure below), as well as an improvement in vascular function and hemodynamics.

PMX Hemoperfusion Therapy

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