In:
International Archives of Allergy and Immunology, S. Karger AG, Vol. 118, No. 2-4 ( 1999), p. 426-428
Abstract:
〈 b 〉 Background: 〈 /b 〉 Eosinophil granule major basic protein (MBP) mediates many eosinophil–associated immune functions and it adheres eosinophils to parasite targets. 〈 b 〉 Methods: 〈 /b 〉 We compared the toxicities of MBP and melittin to K562 and HL–60 cells using five cytotoxicity methods. 〈 b 〉 Results: 〈 /b 〉 Trypan blue staining, propidium iodide/ CellTracker 〈 sup 〉 ™ 〈 /sup 〉 Green staining and incorporation of 〈 sup 〉 14 〈 /sup 〉 C–leucine assays indicated that MBP damages most cells by 1 h. In contrast, 〈 sup 〉 51 〈 /sup 〉 Cr and lactic dehydrogenase (LDH) release assays indicated that MBP damages most cells only at 20 h. All five methods indicated that melittin damages nearly all cells by 1 h. To resolve these discrepancies, the procedures were modified. Without cell transfer, dye staining methods showed that MBP produces very little cytotoxicity at 4 h. 〈 sup 〉 51 〈 /sup 〉 Cr and LDH assays, modified to mimic cell transfer, showed increased cytotoxicities at 4 h. The 〈 sup 〉 14 〈 /sup 〉 C–leucine assay modified by solubilization of cells with SDS and by trichloroacetic acid precipitation showed increased recovery of labeled protein and, thus, lower cytotoxicity, about 50%, at 4 h. 〈 b 〉 Conclusion: 〈 /b 〉 Overall, MBP’s ability to cause molecular and cellular adhesion confounds cytotoxicity measurements. A modified 〈 sup 〉 14 〈 /sup 〉 C–leucine assay overcame MBP’s adhesiveness and provided an accurate measure of cytotoxicity.
Type of Medium:
Online Resource
ISSN:
1018-2438
,
1423-0097
Language:
English
Publisher:
S. Karger AG
Publication Date:
1999
detail.hit.zdb_id:
1482722-0
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