Visualization of the enzyme trimethylamine oxide demethylase in isoelectric focusing gels by an enzyme-specific staining method.

An enzyme-specific staining method for trimethylamine oxide demethylase (TMAO-ase) was developed. Direct visualization could be reached by coupling the reactions of the specific TMAO-ase assay with another reaction step generating as final product a dark-blue formazan. For these purposes 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) as tetrazolium salt and phenazine methosulfate (PMS) as electron transfer substance were used.

Comparison of different methods to produce single-strand DNA for identification of canned tuna by single-strand conformation polymorphism analysis.

By using single-strand conformation polymorphism (SSCP) analysis of three amplicons of the cytochrome b gene obtained by the polymerase chain reaction (PCR) it was possible to differentiate between various species of tunas and bonitos processed as canned fish. Four different techniques were used to produce single-strand DNA (ssDNA): (i) Denaturation of double-strand DNA (dsDNA) by formamide and alkali, (ii) two-step asymmetrical PCR, (iii) one-step asymmetrical PCR, and (iv) exonuclease digestion of the phosphorylated strand of dsDNA. The technique rendering optimal results depended on the type of amplicon (i.