Diaminobenzoic acid method

Samples containing DNA are incubated in 1 A perchloric acid for 10 min. Under these conditions, DNA is depurinated, deoxyribose is released, and converted to cohydroxy levulinylaldehyde. An equal volume of 1.3 M DABA dihydrochloride is 

TABLE 1.1. Detection Limits and Interferences for Protein Assay Methods 

Smith 0.2 pg Acidifiers, Cu chelating, reducing agents, Tris, ammonium sulfate, EDTA, lipids + phospholipids. (Neutral detergents and SDS do not interfere) 

Ninhydrin 0.02 pg Amino acids, amine-containing compounds 

Nitration 5 Pg Triton X-100, phenolic compounds. (Tris, SDS, urea, thiols, reducing agents, glycerol, ammonium sulfate, neutral detergents do not interfere) 

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Detailed analysis using several different methods (Pulse and continuous label with tritiated thymidine, DNA fluorescence per cell using the fluorescent Feulgen assay and flow microfluorography (Van Dilla et al., 1969), and DNA specific fluorescence using the diaminobenzoic acid assay (Kissane and Robins, 1958) reveals that the DNA content of a cell increases in a saltatory fashion, and that the early portion of S-phase is a period of low net DNA synthesis which may be mistaken for G1 if insensitive methods of measurement are used (Klevecz et al., 1975). 

The diphenylamine method can be scaled down to increase sensitivity and fluorescent methods are available using diaminobenzoic acid, diamidino-phenylindole (DAP1), ethidium bromide or propidium iodide (Kissane and Robins, 1958 Hinegardner, 1971 Klotz and Zimm, 1972) which are sensitive down to 0.1 fig DNA. 

Similarly to 3,3 -diaminobenzidine, other aromatic o-diamines also react with selenium(rV) in HCl medium. The o-phenylenediamine method [30,31] is more sensitive than the DAB method. The following reagents have been proposed for selenium N-methyl-o--phenylenediamine (e = 1.9-10 at 346 nm) [32], 2-aminodiphenylenediamine [33], 4-nitro-1,2-diaminobenzene [34], 4,5-diamino-2,6-dimercaptopyrimidine [35], l,2-diamino-4-chlorobenzene [36], 4,5,6-triaminopyrimidine [37], 3,4-diaminobenzoic acid [38], and 2,3-diaminonaphthalene [39,40]. 

As with pyrazines (Section 6.2.7.1), phenazines, the dibenzo derivatives of pyrazine, can also be prepared from the reaction of diamines and dicarbonyl compounds. Chattopadhyaya and co-workers utilized this method to build phenazine systems in the synthesis of [Ru(phen)2dppz] 228, which were used for structure elucidation of nucleic acids without radioactive probes <01JACS3551>. The phenazine component of [Ru(phen)2dppz] was prepared by condensation of 226 and 3,4-diaminobenzoic acid. Diketone complex 226 was prepared by two consecutive ligand displacements from RuQs. 

A sensitive fluorimetric assay for 2-amino-2-deoxy-D-hexoses with increased sensitivity over the Elson-Morgan and Dische-Borenfreund indole methods has been reported. The 2,5-anhydrohexoses produced by nitrous acid deamination of heparin, heparan sulphate, and 2-amino-2-deoxy-D-hexoses have been treated with 3,5-diaminobenzoic acid to give stable derivatives. Neither neutral sugars nor acetamido sugars interfere in the reaction. In a modification of an earlier procedure, equivalent colour complexes were formed by the interaction of 3-methyl 2-benzothiazoline and the 2,5-anhydrohexoses produced by deamination of 2-amino-2-deoxy-D-glucose and 2-amino-2-deoxy-D-galactose. ... 

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