chromomycins and aniline-blue

Identification of parameters predicting assisted reproductive technologies (ARTs) success is a major goal of research in reproduction. Quality of gametes is essential to achieve good quality embryos and increase the success of ARTs. We evaluated two sperm parameters, chromatin maturity and expression of the sperm specific calcium channel CATSPER, in relation to ART outcomes in 206 couples undergoing ARTs. Chromatin maturity was evaluated by Chromomycin A3 (CMA3) for protamination and Aniline Blue (AB) for histone persistence and CATSPER expression by a flow cytometric method. CMA3 positivity and CATSPER expression significantly predicted the attainment of good quality embryos with an OR of 6.6 and 14.3 respectively, whereas AB staining was correlated with fertilization rate. In the subgroup of couples with women ≤35 years, CATSPER also predicted achievement of clinical pregnancy (OR = 4.4). Including CMA3, CATSPER and other parameters affecting ART outcomes (female age, female factor and number of MII oocytes), a model that resulted able to predict good embryo quality with high accuracy was developed. CMA3 staining and CATSPER expression may be considered two applicable tools to predict ART success and useful for couple counseling. This is the first study demonstrating a role of CATSPER expression in embryo development after ARTs programs.

Topics: Adult; Aniline Compounds; Calcium Channels; Chromatin; Chromomycins; Female; Fertilization in Vitro; Flow Cytometry; Gene Expression; Humans; Male; Middle Aged; Reproduction; Reproductive Techniques, Assisted; Spermatozoa; Staining and Labeling; Treatment Outcome; Young Adult

The effect of paternal age on sperm DNA fragmentation and decondensation was determined in a retrospective study involving 1769 patients. TdT (terminal deoxynucleotidyl transferase)-mediated dUDP nick-end labelling (TUNEL) assay was used to assess fragmentation, and DNA decondensation was measured with either chromomycin or aniline blue staining. The impact of atypical forms was also analysed. DNA fragmentation increases with age, but is independent of the percentage of atypical forms. Both staining techniques revealed a negative correlation between the quality of sperm packaging and the percentage of atypical forms. Decondensation increases with increasing age and fragmentation when measured with chromomycin; however, an inverse relationship is observed when testing is performed using aniline blue. These observations are discussed in relation to the specificity of the dyes, the deposition of protamines and the impact of age and reactive oxygen species on protamine cross-linking.

Topics: Adult; Aniline Compounds; Chromomycins; DNA; DNA Fragmentation; Humans; Male; Middle Aged; Paternal Age; Spermatozoa

To consider the relationship between different sperm nuclear maturity tests and in vitro fertilization (IVF) rate, in order to select the most sensitive, specific, and independent factor(s) for prediction of in vitro fertilization.. Infertile couples (101) were randomly selected from IVF candidates referred to Isfahan Fertility and Infertility center. Semen samples were collected on the day of oocyte recovery. Following routine semen analysis, major portion of the semen was prepared for routine IVF insemination and the remaining was used for following sperm nuclear maturity tests: chromomycin A3 (CMA3), aniline blue, sodium dodecyl sulphate (SDS) test, and acridine orange test with or without heat shock (87 degrees C, 5 min). Sperms (200) were evaluated for each test. The results were recorded and analyzed for their correlation to fertilization rate, using correlation coefficient, logistic regression analysis, student t-test, and receiver operating characteristics (ROC) curve.. Among these tests, aniline blue and CMA3, and semen parameters, sperm morphology, and sperm motility showed a significant correlation with fertilization rate. Using logistic regression analysis, sperm morphology and CMA3 were the only independent factors related to in vitro fertilization. ROC curves showed that among above tests, CMA3 is the most specific and sensitive for sperm nuclear maturity.. Among CMA3, aniline blue, SDS test, and acridine orange, CMA3 was the most sensitive and specific test that can be used along with routine semen analysis for more precise prediction of fertilization rate.

Topics: Acridine Orange; Aniline Compounds; Cell Nucleus; Cell Size; Chromomycins; Female; Fertilization in Vitro; Fluorescent Dyes; Humans; Male; Semen; Sodium Dodecyl Sulfate; Sperm Count; Sperm Head; Sperm Motility; Spermatozoa; Staining and Labeling; Statistics as Topic