Serum total protein level, lysozyme activity and natural hemolytic (alternative pathway) complement activity from each of the four fish/ aquarium were performed in duplicate as described in.
Total serum protein concentration was determined using the modified Biuret method. Serum lysozyme activity was determined by the method of Litwack as modified by Sankaran and Gurnani by measuring the lytic activity of the tilapia serum against bacterium Micrococcus lysodeikticus.
Serum alternative complement activity, which is expressed as ACH50 (units/mL) and represents the volume of serum necessary to produce lysis of 50% of the target cells under standard conditions, was adapted from Sunyer and Tort (1995) and modified for use in microtiter plates as described in.