Establishing a stable platform for the measurement of blood endotoxin levels in the dialysis population.
Limulus amoebocyte lysate assay
artefacts
endotoxaemia
endotoxin
haemodialysis
inflammation
Journal
Diagnosis (Berlin, Germany)
ISSN: 2194-802X
Titre abrégé: Diagnosis (Berl)
Pays: Germany
ID NLM: 101654734
Informations de publication
Date de publication:
26 05 2021
26 05 2021
Historique:
received:
18
11
2019
accepted:
02
03
2020
pubmed:
19
4
2020
medline:
16
10
2021
entrez:
19
4
2020
Statut:
epublish
Résumé
Gram-negative lipopolysaccharides are potent inducers of inflammation and have been shown to be present in patients with end-stage kidney disease. There are a variety of different manufacturers and assay types to quantify endotoxin levels; however, there is no standard methodology to demonstrate its presence in plasma. A control group consisting of haemodialysis and non-kidney disease was selected. Five sets of experiments were conducted to try and ascertain the best platform for plasma endotoxin testing. This included: testing of blank tubes; the effects of freezing, thawing and storage on recovery; the effect of different buffers; use of an endpoint assay and comparison of turbidimetric vs. chromogenic kinetic assays. No endotoxin was detected in the blood collection tubes. Freezing and thawing per se did not affect spike recovery rates. However, the sequencing of sample dilution relative to freezing had a significant effect on endotoxin recovery. Buffers increased spike recovery at all levels of dilution. No endotoxin was demonstrated with either the turbidimetric or chromogenic kinetic assay at two different dilutions in the haemodialysis controls. The endpoint assay at a 1:5 dilution did not achieve a valid standard curve. The findings of our study suggest that, when testing plasma samples, either a turbidimetric or chromogenic assay may be used and should be diluted with appropriate buffers to achieve optimal recovery. Studies looking to quantify the presence of plasma endotoxin need to internally validate their assays and specify their validation findings in their results.
Sections du résumé
BACKGROUND
Gram-negative lipopolysaccharides are potent inducers of inflammation and have been shown to be present in patients with end-stage kidney disease. There are a variety of different manufacturers and assay types to quantify endotoxin levels; however, there is no standard methodology to demonstrate its presence in plasma.
METHODS
A control group consisting of haemodialysis and non-kidney disease was selected. Five sets of experiments were conducted to try and ascertain the best platform for plasma endotoxin testing. This included: testing of blank tubes; the effects of freezing, thawing and storage on recovery; the effect of different buffers; use of an endpoint assay and comparison of turbidimetric vs. chromogenic kinetic assays.
RESULTS
No endotoxin was detected in the blood collection tubes. Freezing and thawing per se did not affect spike recovery rates. However, the sequencing of sample dilution relative to freezing had a significant effect on endotoxin recovery. Buffers increased spike recovery at all levels of dilution. No endotoxin was demonstrated with either the turbidimetric or chromogenic kinetic assay at two different dilutions in the haemodialysis controls. The endpoint assay at a 1:5 dilution did not achieve a valid standard curve.
CONCLUSIONS
The findings of our study suggest that, when testing plasma samples, either a turbidimetric or chromogenic assay may be used and should be diluted with appropriate buffers to achieve optimal recovery. Studies looking to quantify the presence of plasma endotoxin need to internally validate their assays and specify their validation findings in their results.
Identifiants
pubmed: 32304297
doi: 10.1515/dx-2019-0088
pii: dx-2019-0088
doi:
Substances chimiques
Endotoxins
0
Types de publication
Journal Article
Research Support, Non-U.S. Gov't
Retracted Publication
Langues
eng
Sous-ensembles de citation
IM
Pagination
249-256Commentaires et corrections
Type : RetractionIn
Informations de copyright
©2020 Shyam Dheda et al., published by De Gruyter, Berlin/Boston.
Références
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