Metabolic Endotoxemia, Feeding Studies and the Use of the Limulus Amebocyte (LAL) Assay; Is It Fit for Purpose?
The Limulus amebocyte assay (LAL) is increasingly used to quantify metabolic endotoxemia (ME), particularly in feeding studies. However, the assay was not intended to assess plasma at levels typically seen in ME. We aimed to optimize and validate the LAL assay under a range of pre-treatment conditio...
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| Format: | Article |
| Language: | English |
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MDPI AG
2020-06-01
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| Series: | Diagnostics |
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| Online Access: | https://www.mdpi.com/2075-4418/10/6/428 |
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| author | Karma Pearce Dianne Estanislao Sinan Fareed Kelton Tremellen |
| author_facet | Karma Pearce Dianne Estanislao Sinan Fareed Kelton Tremellen |
| author_sort | Karma Pearce |
| collection | DOAJ |
| description | The Limulus amebocyte assay (LAL) is increasingly used to quantify metabolic endotoxemia (ME), particularly in feeding studies. However, the assay was not intended to assess plasma at levels typically seen in ME. We aimed to optimize and validate the LAL assay under a range of pre-treatment conditions against the well-established lipopolysaccharide binding protein assay (LBP). Fifteen healthy overweight and obese males aged 28.8 ± 9.1years provided plasma. The LAL assay employed a range of pre-treatments; 70 °C for 15 and 30 min and 80 °C for 15 and 30 min, ultrasonication (70 °C for 10 min and then 40 °C for 10 min), and dilution (1:50, 1:75, 1:100, and 1:200 parts) or diluted using 0.5% pyrosperse. Seventeen different plasma pre-treatment methods employed prior to the use of the LAL analytical technique failed to show any relationships with either LBP, or body mass index (BMI; obesity), the biological trigger for ME (<i>p</i> > 0.05 for all). As expected, BMI positively correlated with LBP (r = 0.523, <i>p</i> = 0.052. No relationships were observed between LAL with any of the sample pre-treatments and LBP or BMI. In its current form, the LAL assay is unsuitable for detecting levels of endotoxin typically seen in ME. |
| first_indexed | 2024-03-10T18:54:56Z |
| format | Article |
| id | doaj.art-14fbde748cf543c6b9c199ee03544f37 |
| institution | Directory Open Access Journal |
| issn | 2075-4418 |
| language | English |
| last_indexed | 2024-03-10T18:54:56Z |
| publishDate | 2020-06-01 |
| publisher | MDPI AG |
| record_format | Article |
| series | Diagnostics |
| spelling | doaj.art-14fbde748cf543c6b9c199ee03544f372023-11-20T04:48:33ZengMDPI AGDiagnostics2075-44182020-06-0110642810.3390/diagnostics10060428Metabolic Endotoxemia, Feeding Studies and the Use of the Limulus Amebocyte (LAL) Assay; Is It Fit for Purpose?Karma Pearce0Dianne Estanislao1Sinan Fareed2Kelton Tremellen3Division of Health Sciences, School of Pharmacy and Medical Sciences & Alliance for Research in Exercise Nutrition and Activity (ARENA), University of South Australia, Adelaide SA 5001, AustraliaDivision of Health Sciences, School of Pharmacy and Medical Sciences & Alliance for Research in Exercise Nutrition and Activity (ARENA), University of South Australia, Adelaide SA 5001, AustraliaDivision of Health Sciences, School of Pharmacy and Medical Sciences & Alliance for Research in Exercise Nutrition and Activity (ARENA), University of South Australia, Adelaide SA 5001, AustraliaDepartment of Obstetrics Gynaecology and Reproductive Medicine, Flinders University, Bedford Park SA 5042, AustraliaThe Limulus amebocyte assay (LAL) is increasingly used to quantify metabolic endotoxemia (ME), particularly in feeding studies. However, the assay was not intended to assess plasma at levels typically seen in ME. We aimed to optimize and validate the LAL assay under a range of pre-treatment conditions against the well-established lipopolysaccharide binding protein assay (LBP). Fifteen healthy overweight and obese males aged 28.8 ± 9.1years provided plasma. The LAL assay employed a range of pre-treatments; 70 °C for 15 and 30 min and 80 °C for 15 and 30 min, ultrasonication (70 °C for 10 min and then 40 °C for 10 min), and dilution (1:50, 1:75, 1:100, and 1:200 parts) or diluted using 0.5% pyrosperse. Seventeen different plasma pre-treatment methods employed prior to the use of the LAL analytical technique failed to show any relationships with either LBP, or body mass index (BMI; obesity), the biological trigger for ME (<i>p</i> > 0.05 for all). As expected, BMI positively correlated with LBP (r = 0.523, <i>p</i> = 0.052. No relationships were observed between LAL with any of the sample pre-treatments and LBP or BMI. In its current form, the LAL assay is unsuitable for detecting levels of endotoxin typically seen in ME.https://www.mdpi.com/2075-4418/10/6/428metabolic endotoxemiaendotoxinslipopolysaccharideslimulus amebocyte (LAL) assaylipopolysaccharide binding protein (LBP) assayadiposity |
| spellingShingle | Karma Pearce Dianne Estanislao Sinan Fareed Kelton Tremellen Metabolic Endotoxemia, Feeding Studies and the Use of the Limulus Amebocyte (LAL) Assay; Is It Fit for Purpose? Diagnostics metabolic endotoxemia endotoxins lipopolysaccharides limulus amebocyte (LAL) assay lipopolysaccharide binding protein (LBP) assay adiposity |
| title | Metabolic Endotoxemia, Feeding Studies and the Use of the Limulus Amebocyte (LAL) Assay; Is It Fit for Purpose? |
| title_full | Metabolic Endotoxemia, Feeding Studies and the Use of the Limulus Amebocyte (LAL) Assay; Is It Fit for Purpose? |
| title_fullStr | Metabolic Endotoxemia, Feeding Studies and the Use of the Limulus Amebocyte (LAL) Assay; Is It Fit for Purpose? |
| title_full_unstemmed | Metabolic Endotoxemia, Feeding Studies and the Use of the Limulus Amebocyte (LAL) Assay; Is It Fit for Purpose? |
| title_short | Metabolic Endotoxemia, Feeding Studies and the Use of the Limulus Amebocyte (LAL) Assay; Is It Fit for Purpose? |
| title_sort | metabolic endotoxemia feeding studies and the use of the limulus amebocyte lal assay is it fit for purpose |
| topic | metabolic endotoxemia endotoxins lipopolysaccharides limulus amebocyte (LAL) assay lipopolysaccharide binding protein (LBP) assay adiposity |
| url | https://www.mdpi.com/2075-4418/10/6/428 |
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