Biology ETDs

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Coagulase production was investigated for 23 strains of Staphylococcus aureus. The coagulase from seven of these strains was purified and subjected to electrophoresis.

The dialyzable fraction of heart infusion broth was the growth medium. Coagulase was harvested from 5 hr shake cul­tures incubated at 37 c.

A series of dialysis steps lasting 24 hr at a tempera­ture of 2 C were used to purify the different coagulases. The crude coagulase was first separated from the growth med­ium by dialysis against deionized water. The coagulase was next precipitated by dialysis against a £H 3.8 acetate buf­fer which had an ionic strength of 0.1. This was followed by another precipitation in a £H 5.2 acetate buffer with an ionic strength of 0.05. The remaining impurities were pre­cipitated by dialysis against a £H 6.1 phosphate buffer with an ionic strength of 0.1. The three preceding buffers were prepared in deionized water containing 10% ethanol by volume. The final preparation was dialyzed against a EH 8.6 barbital buffer with an ionic strength of 0.075.

The seven purified coagulases were characterized by cel­lulose acetate electrophoresis. ·􀀕ach coagulase was electro­phoresed for 110 min at 1.5 ma/cm strip length in a £H 8.6 barbital buffer having an ionic strength of 0.075. One of each set of electropherograms was stained with either Ponceau Sor Nigrosin and a duplicate was placed on fibrinogen agar to determine whether the stained bands represented active coagulase. Active coagulase produced precipitation bands in the agar. A total of 14 bands were demonstrated for the seven coagulases. Most of the purified coagulases were separated into several electrophoretically distinct forms. It was suggested that coagulase is an isoenzyme.



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Department Name

UNM Biology Department

First Committee Member (Chair)

James S. Booth

Second Committee Member

Gordon Johnson

Third Committee Member

John W. Beakley

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