staining: To perform this staining, fresh sperm smear of each case was air dried and then fixed in 3% buffered glutaraldehyde in 0.2 M phosphate buffer (pH=7.2) for 30 min at room temperature.
Furthermore, the use of aniline blue
staining revealed the granulosa cells in the control culture showed the round nuclei of euchromatin in the centre of their cytoplasm.
x [10.sup.6]/ AB+sperm (%) High gr testis) color Control 18.8 [+ or -] .32 68.6 [+ or -] 3.3 Sulfa 15.3 [+ or -] 0.50* 88.1 [+ or -] 1.3 * Sulfa+vit C 23.6 [+ or -] 0.60* 53.5 [+ or -] 4.5 * Sulfa + vit E 28.9 [+ or -] 0.57 (#) 27.8 [+ or -] 3.1 (#) Sulfa + vit E 38.5 [+ or -] 1.5 (#) 11.2 [+ or -] 1.3 (#) & vit C Sulfa: sulfasalazine; vit C: vitamin C; vit E: vitamin E; DSP: daily sperm production (DSP); AB+: Aniline Blue
positive; * Indicate significant difference from the control group; (#) Indicate significant difference from treated with Sulfa alone
Several tests such as TdT (terminal deoxyribonucleotidyl transferase)-mediated dUTP nick-end labeling (TUNEL) assay, aniline blue
straining, chromomycin A3 and acridine orange (AO) staining have been described to determine the extent of DNA damage.
Brewbaker and Kwack (B & K), B & K + aniline blue
, and FCR media were made fresh daily before use.
- Pollen was stained with aniline blue
lactophenol to increase visibility and indicate viability (Kearns and Inouye, 1993).
Staining in decolorized Aniline blue
(DAB) for 24 h was followed by mounting in a drop of DAB: glycerin (1:1) solution.
Currently, different types of cyotchemical tests such as aniline blue
, toluiden blue, chromomycine A3 and acridine orang stainings, sperm chromatin dispersion (SCD), sperm chromatin structure assay (SCSA), comet assay and TUNEL assay are available for sperm DNA and chromatin evaluation and each of them has its own advantages and disadvantages (2).
The Acidic Aniline Blue
(AAB) stain discriminates between lysine-rich histones and arginine/cysteine-rich protamines.
Pollen viability was estimated by utilizing the lactophenol aniline blue
Acidic Aniline Blue
Staining (AABS): During histone replacement with protamines and chromatin condensation in spermatogenesis, normal sperm do not stain with aniline blue
, but sperm with defective density or immature sperm absorb the stain (28).
However, when chromatin integrity was measured by aniline blue
divided in two groups according to a threshold of 10%, the fertilization rate was significantly higher for the group with less than 10% chromatin abnormality(85 [+ or -] 22.3 vs.