Development of a sodium dodecyl sulfate-polyacrylamide gel electrophoresisreference method for the analysis and identification of fish species in raw and heat-processed samples: A collaborative study

A collaborative study was carried out in seven European labs with the aim o f achieving a sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SD S-PAGE) standard operation procedure to identify fish species in raw and co oked samples. Urea and SDS-containing solutions were evaluated as extractan ts. Several preelectrophoretic operations - such as treatment with RNase/DN ase, ultrafiltration and desalting - and up to ten types of gels and three SDS-PAGE systems were considered. The SDS-containing solution allowed a hig her protein extractability than urea. Unlike urea extraction, SDS extractio n seemed not to be influenced so much by the state of the sample (raw, cook ed at 60 degrees C, cooked at 85 degrees C). Desalting, ultrafiltration or treatment with RNase/DNase did not improve the discriminatory power of the protein patterns. Commercial homogeneous 15% ExcelGels, especially when the y were silver stained, yielded good results and afforded higher reproducibi lity, thus allowing a better matching of results among the laboratories par ticipating in this collaborative study. Under the optimized technical condi tions described above, all the fish species tested, either raw and cooked, yielded reproducible and discriminant species-specific protein patterns.