title: Double probing individual human spermatozoa: aniline blue staining for persistent histones and fluorescence in situ hybridization for aneuploidies.
abstract: OBJECTIVE
  To study the potential relationship between two sperm nuclear attributes: persistence of histones and occurrence of chromosomal aneuploidies.
  
  
  METHODS
  The two variables were examined by double probing of the same spermatozoa.
  
  
  METHODS
  Academic Andrology Laboratory.
  
  
  METHODS
  Semen samples subjected for analyses were examined.
  
  
  METHODS
  We studied >58,000 spermatozoa, in seven men, first with aniline blue histone staining, graded as light (mature sperm), intermediate (moderately immature), and dark (severely arrested maturation). After recording the staining patterns and destaining, the same spermatozoa were subjected to fluorescence in situ hybridization (FISH), using centrometric X, Y, and 17 chromosome probes.
  
  
  METHODS
  Proportions of sperm with light, intermediate, and dark staining were assessed, and ploidy of these sperm was evaluated.
  
  
  RESULTS
  The aneuploidy frequencies in intermediate versus light (mature) spermatozoa were increased four- to sixfold. In addition, aneuploidy frequencies and proportions of intermediate sperm were related. There was no FISH signal detectable in the darkly stained, severely arrested mature sperm.
  
  
  CONCLUSIONS
  The data suggest that in sperm with arrested maturity and DNA fragmentation, the binding of FISH probes is diminished. DNA damage is further aggravated by the decondensation and denaturation steps of FISH. Thus, there is a strong likelihood that in oligozoospermic men, with a higher proportion of sperm with arrested maturation, the sperm disomy frequencies are historically underestimated.
authors: Yale Sperm Physiology Laboratory, Yale University, School of Medicine, New Haven, Connecticut 06510, USA.
authors: Laszlo
authors: L
authors: Ovári
authors: Leyla
authors: L
authors: Sati
authors: Jill
authors: J
authors: Stronk
authors: Antal
authors: A
authors: Borsos
authors: David C
authors: DC
authors: Ward
authors: Gabor
authors: G
authors: Huszar
mesh_terms: D000782:Aneuploidy; D000814:Aniline Compounds / Q000494:pharmacology*; D004249:DNA Damage / Q000502:physiology; D005456:Fluorescent Dyes / Q000494:pharmacology; D006657:Histones / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D007248:Infertility, Male / Q000175:diagnosis / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D007621:Karyotyping / Q000379:methods; D008297:Male; D055101:Semen Analysis / Q000379:methods; D013094:Spermatozoa / Q000166:cytology* / Q000378:metabolism; D013194:Staining and Labeling / Q000379:methods*
chemical_list: D000814:Aniline Compounds; D005456:Fluorescent Dyes; D006657:Histones; C017006:aniline blue