All fluorescence observations were made with a Zeiss Standard 14 photomicroscope with epifluorescence system equipped with an automatic C 35 camera loaded either with Kodak Gold 100 ASA film, for slides stained with acridine
orange (AO) and ethidium bromide (EB), or with Kodak Ultra 400 ASA, for slides strained with JC-1 and Annexin V-FITC.
The test of fluorescence with acridine
orange, based on the determination of the relation between green orthocromatic fluorescence (normal) and red metachromatic (altered) chromatin, is an adequate test to measure effects of chemical agents on male gametes (Andreetta et al., 1995; Evenson et al., 2002).
The effect of washing, frequency of washing, heating of tampon solution and concentration of acridine
orange was compared using non parametric Mann-Whitney test.
Moreover, fluorescence microscopy employing double staining with Acridine
Orange/Bromide Ethidium (OA/BE) (Figure 3) revealed that no significant cell death was observed in the untreated control group (medium), whereas all types of propolis extract (50 [micro]g/mL), as well as the positive control (Amph.
Propodium Iodide (PI) and Acridine
Orange (AO) were obtained from Sigma (USA).
Biofilms were incubated for 24 h and stained with acridine
orange accumulation in acid organelles of normal and vacuolated frog skeletal muscle fibres," Cell Biology International, vol.
Thus, the sensitivity and specificity of these antibodies are clearly maintained after the use of BABB as a clearing agent and SYTOX Green or acridine
orange as a fluorescent nuclear stain.
Dimethyl sulfoxide (DMSO), fetal bovine serum (FBS), penicillin-streptomycin, propidium iodide, acridine
orange, trypsin-EDTA, and MTT powder were purchased from Sigma Chemical Co.
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).
In previous reports, different dyes such as Hoechst 33258 , acridine
orange , thiazole orange , or hydroethidine  were used to determine parasitemia in cultures of Plasmodium falciparum by FACS.
Cells were stained with 10-n-nonyl acridine
orange (NAO) for 30 min, washed with PBS three times and fixed with 1 per cent paraformaldehyde at 48 or 72 h post-irradiation and were analyzed by flow cytometry (23).