Automated electrokinetic analysis; description and application in virology and cell biology.
The electrophoretic mobility (EPM) of selected macromolecules in solution was shown to be accurately determined using an automated electrokinetic analyzer, the PenKem S3000. In addition, the S3000 was used to monitor the effects of T4 phage infection on the EPM of Escherichia coli B. EPM, expressed as the ratio of velocity in microns/sec to field strength in V/cm, was measured for calf thymus DNA, for pneumococcal capsular polysaccharide serotype 3 (PCP-3), and for bovine serum albumin (BSA) unbound in solution; values of -3.05, -2.736 and -1.176, respectively, were obtained. The EPM of these macromolecules remained the same when they were bound to latex beads. The S3000 may therefore be suitable for measurement of the EPM of unbound macromolecules. The EPM of T4 phage in solution was measured to be -1.203. However, both the zwitterionic latex-bound T4 phage as well as T4 phage disrupted by ultrasonication exhibited an EPM of approximately -2.50, suggesting to us that binding to zwitterionic latex may cause release of phage DNA. The notion that phage DNA is responsible for the increased negative charge was supported by the observation that the EPM of E. coli B increased to the level of free DNA within 5 min when E. coli B (the host cell for phage T4) had been exposed to 10 phage particles per cell. Electronmicrographs of phage infected E. coli B cells showed numerous strands of free DNA at the bacterial surface. It is concluded that the S3000 not only measures the EPM of macromolecules in solution but that the instrument can be used also to monitor the behavior of the host cell surface in response to attachment of viral particles.
Sloyer
J L
JL
Institute for Cancer Research, Fox Chase Cancer Center, Philadelphia, Pennsylvania.
Bayer
M E
ME
D001322:Autoanalysis; D002448:Cell Adhesion; D004247:DNA / Q000648:ultrastructure*; D004563:Electrochemistry; D004926:Escherichia coli / Q000235:genetics / Q000648:ultrastructure; D007700:Kinetics; D013604:T-Phages / Q000235:genetics / Q000648:ultrastructure; D014777:Virus Diseases / Q000175:diagnosis*
D004247:DNA