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Mutagenesis, Vol. 16, No. 6, 455-459, November 2001
© 2001 UK Environmental Mutagen Society/Oxford University Press

Lysis of whole blood in vitro causes DNA strand breaks in human lymphocytes

S. Narayanan2, M.R. O'Donovan1 and S.J. Duthie2,2

2 Rowett Research Institute, Greenburn Road, Bucksburn, Aberdeen AB21 9SB and 1 Safety Assessment, AstraZeneca R&D, Bakewell Road, Loughborough LE11 5RH, UK

DNA damage in lymphocytes, as measured by alkaline single cell gel electrophoresis (pH 12.7), is greatly increased by the concurrent lysis of whole blood in both freshly isolated samples and in PHA-stimulated cultures over a period of 7 days. Further, there is a marked progressive increase in DNA damage with time in PHA-stimulated lymphocytes cultured in whole blood even when the lymphocytes are separated before analysis; no such increase is seen in lymphocytes cultured alone. This indicates that there are components in whole blood that can cause DNA damage in lymphocytes, with granulocytes and lysis of red blood cells likely candidates. The DNA damage is greatly reduced in granulocyte-depleted whole blood cultures, but even in these significant increases are seen at later sampling times. Consequently, careful sample preparation is of paramount importance if the Comet assay is to be successfully used to assess DNA damage in human peripheral blood lymphocytes. Further, the progressive increase in DNA damage in whole blood cultures may influence other methods using lymphocytes for population biomonitoring and may be significant for in vitro genotoxicity testing.

2 To whom correspondence should be addressed. Tel: +44 1224 712751; Fax: +44 1224 716629; Email: sd{at}rri.sari.ac.uk


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