Reduction in signal overlap results in increased FISH efficiency: implications for preimplantation genetic diagnosis.
BACKGROUND
In the absence of mosaicism, one of the problems of preimplantation genetic diagnosis with FISH is the occurrence of false-negative hybridization results. It has been hypothesized that missing signals are produced by spatial overlap of signals.


RESULTS
To investigate the relation among cell density, signal overlap, and hybridization signal detection, 371 blastomeres and 4556 lymphocytes were fixed in different cellular concentrations and analyzed by FISH using probes for chromosomes X, Y, and 18, and their nuclear diameters and FISH results scored. The results showed that the lower the diameter of fixed nuclei, the higher the number of signal overlaps and missing signals. The minimum number of missing signals was obtained when lymphocyte and blastomere nuclei had 40 or more microns in diameter after fixation and FISH. Since blastomeres were fixed individually, results with blastomeres were invariably better than with lymphocytes.
Munné
S
S
New York Hospital-Cornell University Medical Center, New York, USA.
Dailey
T
T
Finkelstein
M
M
Weier
H U
HU
D000782:Aneuploidy; D016477:Artifacts; D001757:Blastomeres / Q000648:ultrastructure*; D002452:Cell Count; D002467:Cell Nucleus / Q000648:ultrastructure*; D048429:Cell Size; D002869:Chromosome Aberrations / Q000175:diagnosis* / Q000196:embryology / Q000235:genetics / Q000517:prevention & control; D025063:Chromosome Disorders; D015342:DNA Probes; D005188:False Negative Reactions; D005260:Female; D005315:Fetal Diseases / Q000175:diagnosis* / Q000235:genetics / Q000517:prevention & control; D030342:Genetic Diseases, Inborn / Q000175:diagnosis* / Q000196:embryology / Q000235:genetics / Q000517:prevention & control; D008040:Genetic Linkage; D005819:Genetic Markers; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D008214:Lymphocytes / Q000648:ultrastructure*; D008297:Male; D011296:Prenatal Diagnosis / Q000379:methods*; D012732:Sex Determination Analysis / Q000379:methods*; D016707:Tissue Fixation / Q000379:methods*; D014960:X Chromosome / Q000235:genetics*
D015342:DNA Probes; D005819:Genetic Markers