Quantification of Defensive Proteins in Skin Mucus of Using Minimally Invasive Sampling and High-Sensitivity ELISA.

Protocols used to collect fish skin mucus may inadvertently compromise the sampled fish or the resulting sample. Here, we used three methods (wiping, scraping, and absorption) to collect skin mucus from Atlantic salmon and compared their invasiveness on fish skin epithelium. We found that the absorption method was the least invasive. We also compared the abundance of antigen-specific immunoglobulin M subtype A antibodies (IgM-A Ab) and complement component 5 (C5) in mucus samples collected from vaccinated fish by the three methods. An enzyme-cascade-amplification strategy colorimetric immune assay was optimized and used to analyze IgM-A, and ELISA was used to analyze C5. The abundance of antigen-specific IgM-A in skin mucus was comparable between the three methods, but C5 was significantly lower in absorbed mucus in comparison to in the wiped or scraped mucus samples. Absorbed skin mucus samples collected from various body regions of salmon, levels of C5 were comparable, while specific IgM-A amounts varied between the regions. By comparing three mucus-absorbing materials (medical wipe, gauze, and cotton) for their ability to absorb and release IgM-A and C5, medical wipes proved to be ideal for IgM-A analysis, whereas gauze was the best for C5 analysis.