“Dirty little secrets”—Endotoxin contamination of recombinant proteins

Wakelin, Sonia J.; Sabroe, Ian; Gregory, Christopher D.; Poxton, Ian R.; Forsythe, John; Garden, O. James; Howie, Sarah

doi:10.1016/j.imlet.2006.04.007

Cited by 92 publications

(70 citation statements)

References 87 publications

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“…The phenomenon has recently been addressed by the German Hyglos company 9 , a new kid on the block of pyrogen testing. For recombinant proteins, this problem has been nicely addressed by Wakelin et al (2006), showing that whenever an immune mediator or other protein shows endotoxin-like properties, these probably stem from an endotoxin contamination. Foster Jordan, the CEO of Charles-River Endosafe, the largest LAL manufacturer, once told me that 90% of LAL testing is done on water samples.…”

Section: Cryo-blood Assaysmentioning

confidence: 99%

“…Pretesting of clinical grade erythrocytes and thrombocytes intended for transfusion indicated interference-free retrieval of an endotoxin spike (Schindler et al, 2009) when compared to the saline control. Possible masking of pyrogens binding to cells or remaining plasma proteins is a special concern for the LAL (Wakelin et al, 2006). Noteworthy, the whole blood MAT has been show to handle this especially well (Spreitzer et al, 2002;PerdomoMorales et al, 2011).…”

Section: Slow Implementation Of the Pyrogen Test For Medical Devicesmentioning

confidence: 99%

See 1 more Smart Citation

The human whole blood pyrogen test – lessons learned in twenty years

Hartung

2015

ALTEX

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Section: Cryo-blood Assaysmentioning

confidence: 99%

Section: Slow Implementation Of the Pyrogen Test For Medical Devicesmentioning

confidence: 99%

The human whole blood pyrogen test – lessons learned in twenty years

Hartung

2015

ALTEX

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“…Endotoxin (such as LPS), a putative stimulator of acute inflammation, is a common contaminant in protein preparations (46,47). Endotoxin levels were determined in the reagents we tested for inflammatory activity in the air pouch model, along with a summary of their activity to stimulate the innate immune response (Table I).…”

Section: Endotoxin In Bovine Testis-derived Type I-s and Iv-s Hyals Smentioning

confidence: 99%

Recombinant Human Hyaluronidase PH20 Does Not Stimulate an Acute Inflammatory Response and Inhibits Lipopolysaccharide-Induced Neutrophil Recruitment in the Air Pouch Model of Inflammation

Huang

Zhao

Chen

et al. 2014

The Journal of Immunology

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Hyaluronidase (Hyal) and low m.w. hyaluronan (LMW HA) fragments have been widely reported to stimulate the innate immune response. However, most hyaluronidases used were purified from animal tissues (e.g., bovine testis Hyal [BTH]), and contain endotoxin and other unrelated proteins. We tested a highly purified recombinant human Hyal (rHuPH20) and endotoxin-free HA fragments from Mr 5,000 to 1,500,000 in the rodent air pouch model of inflammation to determine their potential for stimulation of the innate immune response. Exogenous LMW HA fragments (average Mr 200,000) failed to induce either cytokine/chemokine production or neutrophil infiltration into the air pouch. Challenging the air pouch with LPS or BTH stimulated production of cytokines and chemokines but rHuPH20 did not, suggesting that neither PH20 nor generation of LMW HA fragments in situ stimulates cytokine and chemokine production. LPS and BTH also induced neutrophil infiltration into the air pouch, which was not observed with rHuPH20 treatment. Endotoxin-depleted BTH had much reduced proinflammatory activity, suggesting that the difference in inflammatory responses between rHuPH20 and BTH is likely due to endotoxin contaminants in BTH. When rHuPH20 was dosed with LPS, the induction of cytokines and chemokines was the same as LPS alone, but neutrophil infiltration was inhibited, likely by interrupting HA–CD44 interaction. Our results indicate that neither rHuPH20 nor its directly generated HA catabolites have inflammatory properties in the air pouch model, and rHuPH20 can instead inhibit some aspects of inflammation, such as neutrophil infiltration into the air pouch.

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“…the enormous potency of endotoxins (bacterial toxins are designed by evolution to damage) also might lead to another often-overlooked phenomenon: we rarely test for endotoxin contaminations of test substances. For example, recombinant proteins can absorb endotoxins that are then difficult to trace or remove (Wakelin et al, 2006). Similarly, nanoparticles can carry pyrogenic contaminations that are difficult to detect but biologically highly active (Jones and Grainger, 2009).…”

Section: Considerationmentioning

confidence: 99%