Measuring Total Classical Pathway and Activities of Individual Components of the Mouse Complement Pathway

Abstract

The complement system is a central component of innate immunity, responsible for recognition and killing of bacteria by tagging invaders through opsonisation, thereby promoting phagocytosis, and by direct lysis. Complement activity is routinely measured using functional assays that utilise erythrocytes as targets. The classical pathway haemolytic assay (CH50) with antibody sensitised sheep erythrocytes as target is used worldwide in clinical and research laboratories to measure complement activity in human and rodent sera. While there are no particular limitations in the human assay, measuring complement in mouse serum is more difficult and usually requires large amounts of serum, which is challenging to collect in experiments. In particular, it is challenging to measure the activities of individual mouse complement proteins. To overcome this hurdle, we have developed protocols that employ human sera depleted of single complement proteins as the source of the other complement proteins and test mouse serum to restore the relevant component. This simple haemolytic assay is a useful tool for confirming natural or engineered complement deficiencies and complement dysregulation in mouse models.