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The fifth component of complement (C5): purification without activation.

A Dessauer, U Rother

    Immunobiology
    |May 1, 1983
    PubMed
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    Researchers developed a new method to purify complement component 5 (C5), preventing its activation during the process. This ensures C5 is not prematurely activated, improving complement system studies.

    Area of Science:

    • Immunology
    • Biochemistry
    • Complement System Research

    Background:

    • Previous purification methods for complement component 5 (C5) often resulted in its partial activation, forming C56.
    • Activated C56 could spontaneously mediate cell lysis with complement components C7-C9, complicating functional studies.
    • Physicochemical conditions like extreme pH or high ionic strength were identified as triggers for C5 activation during purification.

    Purpose of the Study:

    • To develop a novel purification protocol for complement component 5 (C5) that avoids premature activation.
    • To obtain a C5 preparation free from activated C56, enabling accurate downstream functional assays.
    • To confirm that the new C5 preparation requires complement component 6 (C6) for activation.

    Main Methods:

    Related Experiment Videos

  • A modified purification strategy was employed, specifically avoiding polyethyleneglycol (PEG) or ammonium sulfate precipitation.
  • Conditions involving low or high pH were excluded during the purification process.
  • Affinity chromatography was utilized as a final step to remove residual complement component 6 (C6).

    • Main Results:

    • The developed purification method successfully yielded C5 preparations free of activated C56.
    • The purified C5 did not spontaneously mediate cell lysis, unlike preparations from previous methods.
    • Acidification of the new C5 preparation did not induce lysis without the addition of complement component 6 (C6).

    Conclusions:

    • The novel purification method effectively prevents C5 activation, yielding a stable and functional C5 preparation.
    • This improved C5 preparation is suitable for accurate studies of the complement system's lytic pathway.
    • The method ensures the integrity of C5, allowing for precise investigation of its role in immune responses.