Mechanism of connective tissue techniques I. The effect of dye concentration and staining time on anionic dye procedures
Tóm tắt
Anionic dye connective tissue procedures were performed by staining for 5 min and 24 h with (a) 0.00018m and 0.0018m solutions of 28 dyes, and 0.018m solutions of 21 dyes in saturated picric acid (SPA), and (b) 0.0018m and 0.018m solutions of 20 dyes in 1% (w/v) phosphomolybdic acid (PMA). The staining obtained with dyes in SPA was classified as selective (no cytoplasmic staining), moderately selective (traces of cytoplasmic staining) and non-selective (all other staining patterns). The staining of collagen and cytoplasm with dyes in PMA was separately classified on a scale of 1–5 (1 = no staining, 5 = maximum staining). The selectivity of the staining obtained with SPA with solutions of dyes at concentrations of 0.00018m and 0.0018m, and both staining times, was correlated (p < 0.001) with an empirical sulphonic acid constant (SAC) defined as the (number of dye sulphonic acid groups/dye molecular weight) × 103. Correlation with molecular weight was poor and was significant only when staining was performed with 0.00018m dye solutions for 24 h. The dyes were divisible into three groups: group 1 (selectivity independent, or almost independent of staining time), group 2 (selective to moderately selective when staining was performed for 5 min), and group 3 (non-selective). The SAC of the group 1 dyes differed significantly from those of the group 2 and 3 dyes. Selectivity was essentially lost at dye concentrations of 0.018m. The staining with acidic dyes (no amines or substituted amines) in PMA differed significantly (p < 0.001) from that obtained with amphoteric dyes (containing basic substituents). In general, acidic dyes stained cytoplasm. Amphoteric dyes with the exception of indigocarmine stained collagen. However, most of these dyes also stained cytoplasm. In contrast to the results obtained with dyes in SPA, selectivity correlated strongly with molecular weight and only poorly with the SAC. Staining time and dye concentration affected selectivity only when the acidic dyes were used for 5 min at concentrations of 0.0018m and 0.018m. The data obtained do not permit a clear distinction between the rate control and chemical affinity models for the mechanism of staining with anionic dyes. However, it seems possible that different groups of dyes stain by different mechanisms.
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