Partial purification of human leukocytic pyrogen

Area of Science:

• Immunology
• Biochemistry
• Molecular Biology

Background:

• Human leukocytes release leukocytic pyrogen (LP), a protein mediating fever.
• Studying LP requires isolating it from complex mixtures of proteins in leukocyte supernates.

Purpose of the Study:

• To develop reliable techniques for the initial purification of human leukocytic pyrogen (LP).
• To characterize LP during purification procedures.

Main Methods:

• In vitro stimulation of human peripheral leukocytes with killed staphylococci.
• Isoelectric focusing, alcohol precipitation, ion-exchange chromatography, and gel-filtration for protein isolation.
• Analysis of molecular weight and isoelectric point during purification.

Main Results:

• Crude leukocyte supernates contained two molecular species of LP, separable by alcohol precipitation.
• Ion-exchange chromatography and gel-filtration yielded partially purified LP with increased specific activity.
• Combined purification methods resulted in LP preparations with 5-6 contaminating proteins.

Conclusions:

• Human LP can be partially purified from complex leukocyte-derived protein mixtures.
• Purification procedures did not alter LP's molecular weight or isoelectric point.
• Established techniques enable reliable initial purification of human LP for further study.