A simple assay for a human serum phospholipase A2 that is associated with high-density lipoproteins
Phospholipase A2 (PLA2) activity is usually assayed with expensive radioactive or chromogenic substrates unsuitable for performing large numbers of assays. We have designed a simple microplate assay for human serum PLA2 using the chromogenic substrate 4-nitro-3-octanoyloxybenzoic acid. Using this su...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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Elsevier
2001-10-01
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| Series: | Journal of Lipid Research |
| Subjects: | |
| Online Access: | http://www.sciencedirect.com/science/article/pii/S0022227520322264 |
| _version_ | 1818899012238966784 |
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| author | Nenad Petrovic Carolyn Grove Paul E. Langton Neil L.A. Misso Philip J. Thompson |
| author_facet | Nenad Petrovic Carolyn Grove Paul E. Langton Neil L.A. Misso Philip J. Thompson |
| author_sort | Nenad Petrovic |
| collection | DOAJ |
| description | Phospholipase A2 (PLA2) activity is usually assayed with expensive radioactive or chromogenic substrates unsuitable for performing large numbers of assays. We have designed a simple microplate assay for human serum PLA2 using the chromogenic substrate 4-nitro-3-octanoyloxybenzoic acid. Using this substrate, serum PLA2 activity was similar to that measured with the previously characterized chromogenic phospholipid substrate 1,2-bis-heptanoylthioglycerophosphocholine. However, the assay described here appears to be more sensitive. The mean PLA2 activity in serum from healthy volunteers (n = 30) measured by this assay was 10.4 ± 1.6 μmol · h−1 · ml−1. The assay is reproducible and is suitable for the analysis of large numbers of samples in a clinical setting. We have also demonstrated that 94% of the PLA2 activity in normal human serum is associated with high-density lipoproteins and that serum PLA2 activity is positively correlated with the lipoprotein parameters total triglyceride (P < 0.0001), total cholesterol (P < 0.0001), and atherogenic index (P = 0.008). The serum PLA2 activity was calcium dependent and was inhibited by the serine protease inhibitor 3,4-dichloroisocoumarin (EC50 = 0.4 mM).The PLA2 activity characterized here is unlikely to be due to plasma platelet-activating factor acetylhydrolase or low molecular weight His-Asp sPLA2, and may represent a new sPLA2 type. |
| first_indexed | 2024-12-19T19:41:12Z |
| format | Article |
| id | doaj.art-2f678a61b4344a69b6fdbfdde3278097 |
| institution | Directory Open Access Journal |
| issn | 0022-2275 |
| language | English |
| last_indexed | 2024-12-19T19:41:12Z |
| publishDate | 2001-10-01 |
| publisher | Elsevier |
| record_format | Article |
| series | Journal of Lipid Research |
| spelling | doaj.art-2f678a61b4344a69b6fdbfdde32780972022-12-21T20:08:14ZengElsevierJournal of Lipid Research0022-22752001-10-01421017061713A simple assay for a human serum phospholipase A2 that is associated with high-density lipoproteinsNenad Petrovic0Carolyn Grove1Paul E. Langton2Neil L.A. Misso3Philip J. Thompson4Asthma and Allergy Research Institute, Ground Floor, E Block, Sir Charles Gairdner Hospital, Nedlands WA 6009, Australia; Department of Medicine, The University of Western Australia, Perth WA 6009, AustraliaAsthma and Allergy Research Institute, Ground Floor, E Block, Sir Charles Gairdner Hospital, Nedlands WA 6009, Australia; Department of Medicine, The University of Western Australia, Perth WA 6009, AustraliaAsthma and Allergy Research Institute, Ground Floor, E Block, Sir Charles Gairdner Hospital, Nedlands WA 6009, Australia; Department of Medicine, The University of Western Australia, Perth WA 6009, AustraliaAsthma and Allergy Research Institute, Ground Floor, E Block, Sir Charles Gairdner Hospital, Nedlands WA 6009, Australia; Department of Medicine, The University of Western Australia, Perth WA 6009, AustraliaAsthma and Allergy Research Institute, Ground Floor, E Block, Sir Charles Gairdner Hospital, Nedlands WA 6009, Australia; Department of Medicine, The University of Western Australia, Perth WA 6009, Australia; To whom correspondence should be addressed. e-mail:Phospholipase A2 (PLA2) activity is usually assayed with expensive radioactive or chromogenic substrates unsuitable for performing large numbers of assays. We have designed a simple microplate assay for human serum PLA2 using the chromogenic substrate 4-nitro-3-octanoyloxybenzoic acid. Using this substrate, serum PLA2 activity was similar to that measured with the previously characterized chromogenic phospholipid substrate 1,2-bis-heptanoylthioglycerophosphocholine. However, the assay described here appears to be more sensitive. The mean PLA2 activity in serum from healthy volunteers (n = 30) measured by this assay was 10.4 ± 1.6 μmol · h−1 · ml−1. The assay is reproducible and is suitable for the analysis of large numbers of samples in a clinical setting. We have also demonstrated that 94% of the PLA2 activity in normal human serum is associated with high-density lipoproteins and that serum PLA2 activity is positively correlated with the lipoprotein parameters total triglyceride (P < 0.0001), total cholesterol (P < 0.0001), and atherogenic index (P = 0.008). The serum PLA2 activity was calcium dependent and was inhibited by the serine protease inhibitor 3,4-dichloroisocoumarin (EC50 = 0.4 mM).The PLA2 activity characterized here is unlikely to be due to plasma platelet-activating factor acetylhydrolase or low molecular weight His-Asp sPLA2, and may represent a new sPLA2 type.http://www.sciencedirect.com/science/article/pii/S0022227520322264enzymesubstratecholesterolatherogenic |
| spellingShingle | Nenad Petrovic Carolyn Grove Paul E. Langton Neil L.A. Misso Philip J. Thompson A simple assay for a human serum phospholipase A2 that is associated with high-density lipoproteins Journal of Lipid Research enzyme substrate cholesterol atherogenic |
| title | A simple assay for a human serum phospholipase A2 that is associated with high-density lipoproteins |
| title_full | A simple assay for a human serum phospholipase A2 that is associated with high-density lipoproteins |
| title_fullStr | A simple assay for a human serum phospholipase A2 that is associated with high-density lipoproteins |
| title_full_unstemmed | A simple assay for a human serum phospholipase A2 that is associated with high-density lipoproteins |
| title_short | A simple assay for a human serum phospholipase A2 that is associated with high-density lipoproteins |
| title_sort | simple assay for a human serum phospholipase a2 that is associated with high density lipoproteins |
| topic | enzyme substrate cholesterol atherogenic |
| url | http://www.sciencedirect.com/science/article/pii/S0022227520322264 |
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