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DNA released from broken cells is often the cause of clumping. DNase I
needs at least 1mM magnesium in order to work effectively, with optimal
activity at about 5mM. The FBS added to the wash buffer would contain some
Mg but perhaps not enough, particularly in the presence of PBS that will
complex divalent cations. Also, actin released from dead cells irreversably
inhibits DNase I.
Try adding some MgCl2 to the wash buffer, but not too much because it will
precipitate, and perhaps increase the concentration DNase in the buffer
when dealing with larger preparations.
I can't think of any other solution, possibly somebody else has a simpler
explanation.
Dean R. Hewish, Cell Biologist & Flow Cytometrist.
CSIRO Biomolecular Engineering, 343 Royal Parade, Parkville, 3052,
Victoria Australia.
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