We. Richards et al., EVALUATING SEX-CHROMOSOME CONTENT OF SORTED HUMAN SPERM SAMPLES WITH USE OF DUAL-COLOR FLUORESCENCE IN-SITU HYBRIDIZATION, American journal of obstetrics and gynecology, 176(6), 1997, pp. 1172-1178
OBJECTIVE: Although most methods for selecting the sex of offspring by
sorting spermatozoa are ineffective at shifting the ratio of Y- to X-
containing cells, some commercial sources continue to offer such servi
ces. Our objective was to evaluate commercially ''sorted'' samples wit
h use of dual-color fluorescence in situ hybridization and to identify
variations in assessment by comparing motile and total sperm populati
ons, donors, observers, and fluorescence in situ hybridization probes.
STUDY DESIGN: Cryopreserved sperm from seven anonymous donors were pr
ocessed as for insemination. Sperm cells from each total sample or mot
ile subfraction were prepared for fluorescence in situ hybridization b
y incubation with disulfide-reducing agents to expand sperm nuclei. Tw
o sets of X and Y chromosome-specific, fluorophore-labeled deoxyribonu
cleic acid probes were used. At least 400 nuclei from each preparation
were classified independently by three blinded observers. Hybridizati
on efficiency, aneuploidy, and sex chromosome content were evaluated i
n subsets of five unsorted, five female-oriented, and five male-orient
ed samples. Total and motile subfractions were compared with eight sam
ples. Fluorescence in situ hybridization probes were compared in five
paired unsorted samples. RESULTS: No differences were detected between
washed samples and paired motile subfractions. No differences in hybr
idization and aneuploidy were detected between groups of sorted sample
s. The Y/X ratio was significantly different between the sorted groups
. However, male-oriented samples had a lower Y/X ratio than female-ori
ented samples did. Observer and probe choice accounted for small but s
ignificant variations that did not alter conclusions about the Y/X rat
io for sorted samples. CONCLUSION: In a series of 10 sorted samples fr
om one commercial source, dual-color fluorescence in situ hybridizatio
n demonstrated a small but significant shift in the sex chromosome rat
ios among samples. However, this shift was opposite to that expected b
y the orientation of the sorted samples.