title: FISH and tips: a large scale analysis of automated versus manual scoring for sperm aneuploidy detection.
abstract: BACKGROUND
  Approximately 1% of the spermatozoa found in ejaculate of healthy men are aneuploid and this rate increases in the population of subfertile and infertile men. Moreover, fertilization with these aneuploid sperm can lead to impaired embryo development. Fluorescent In Situ Hybridization (FISH) is the common cytogenetic tool used for aneuploidy screening on sperm. However, it is a time-consuming technique and cytogenetic or in vitro fertilization laboratories cannot routinely use it and face the increasing demand of such analyses before Assisted Reproductive Techniques (ART). As automation can be a clue for routine practice, this study compares manual and automated scoring of sperm aneuploidy rates using a Metafer Metasystems® device. The results obtained also contribute to global data about FISH on sperm cells.
  
  
  METHODS
  We recruited 100 men addressed for sperm cryopreservation. They all signed an informed consent to participate in the study. 29 men were donors or consulted before vasectomy (control group) and 71 were suffering of Hodgkin's disease or non Hodgkin lymphoma (patient group). One semen sample was collected for each patient, analyzed according to WHO criteria and prepared for a triple-color FISH using centromeric probes for chromosomes 18, X and Y. Automated scoring was performed using a Metafer Metasystems® device.
  
  
  RESULTS
  507,019 cells were scored. We found a strong concordance between the automated and the manual reading (d < 0.01 in Bland-Altman test). We also did not find a statistically significant difference between the automated and the manual reading using Wilcoxon test for total aneuploidy rate (p = 0.06), sex chromosomes disomy (p = 0.33), chromosome 18 disomy (p = 0.39) and diploidy (p = 0.21). Cumulative rate of total aneuploidy was 0.78% ± 0.212% for patient group and 0.54% ± 0.15 for control group and among this, sex chromosome XY disomy rate was of 0.54% for patient group and 0.27% for control group.
  
  
  CONCLUSIONS
  This study validates the automated reading for FISH on sperm with a Metafer Metasystems® device and allows its use in a laboratory routine.
authors.affiliation: Genetic and procreation Department, CS 10217, CHU de Grenoble, Laboratoire d'aide à la procréation - CECOS, Grenoble, Cedex 9, 38043 France ; Joseph Fourier University, Grenoble, F-38000 France ; Laboratoire AGIM, CNRS FRE3405, "Equipe Andrologie Génétique et Cycle cellulaire", La Tronche, F-38700 France.
authors.forename: Guillaume
authors.initials: G
authors.lastname: Martinez
authors.affiliation: Joseph Fourier University, Grenoble, F-38000 France ; ThEMAS TIMC-IMAG, UMR CNRS 5525, Joseph Fourier University, Public Health Pole, University Hospital of Grenoble, BP217, Grenoble, Cedex 9, 38043 France.
authors.forename: Pierre
authors.initials: P
authors.lastname: Gillois
authors.affiliation: Genetic and procreation Department, CS 10217, CHU de Grenoble, Laboratoire d'aide à la procréation - CECOS, Grenoble, Cedex 9, 38043 France ; Joseph Fourier University, Grenoble, F-38000 France.
authors.forename: Marine
authors.initials: M
authors.lastname: Le Mitouard
authors.affiliation: Genetic and procreation Department, CS 10217, CHU de Grenoble, Laboratoire d'aide à la procréation - CECOS, Grenoble, Cedex 9, 38043 France ; Joseph Fourier University, Grenoble, F-38000 France.
authors.forename: Rémy
authors.initials: R
authors.lastname: Borye
authors.affiliation: CECOS and Toulouse University, UPS, Groupe de recherche en fertilité humaine (EA3694, Human Fertility Research Group), Hôpital Paule de Viguier, University Hospital of Toulouse, Toulouse, France.
authors.forename: Camille
authors.initials: C
authors.lastname: Esquerré-Lamare
authors.affiliation: Joseph Fourier University, Grenoble, F-38000 France ; Laboratoire AGIM, CNRS FRE3405, "Equipe Andrologie Génétique et Cycle cellulaire", La Tronche, F-38700 France ; Genetic and Procreation Department, CS10217, CHU de Grenoble, Génétique chromosomique, Grenoble, Cedex 9, 38043 France.
authors.forename: Véronique
authors.initials: V
authors.lastname: Satre
authors.affiliation: CECOS and Toulouse University, UPS, Groupe de recherche en fertilité humaine (EA3694, Human Fertility Research Group), Hôpital Paule de Viguier, University Hospital of Toulouse, Toulouse, France.
authors.forename: Louis
authors.initials: L
authors.lastname: Bujan
authors.affiliation: Genetic and procreation Department, CS 10217, CHU de Grenoble, Laboratoire d'aide à la procréation - CECOS, Grenoble, Cedex 9, 38043 France ; Joseph Fourier University, Grenoble, F-38000 France ; Laboratoire AGIM, CNRS FRE3405, "Equipe Andrologie Génétique et Cycle cellulaire", La Tronche, F-38700 France.
authors.forename: Sylviane
authors.initials: S
authors.lastname: Hennebicq
keywords: Aneuploidy; Automation, Laboratory; In Situ Hybridization, Fluorescence; Infertility; Spermatozoa