High resolution DNA content measurements of mammalian sperm

Journal Article · Thu Dec 31 23:00:00 EST 1981 · Cytometry (Baltimore); (United States)

DOI:https://doi.org/10.1002/cyto.990030103· OSTI ID:6253737

Pinkel, D; Lake, S; Gledhill, B L; Van Dilla, M A; Stephenson, D; Watchmaker, G

The high condensation and flat shape of the mammalian sperm nucleus present unique difficulties to flow cytometric measurement of DNA content. Chromatin compactness makes quantitative fluorescent staining for DNA difficult and causes a high index of refraction. The refractive index makes optical measurements sensitive to sperm head orientation. We demonstrate that the optical problems can be overcome using the commercial ICP22 epiillumination flow cytometer (Ortho Instruments, Westwood, MA) or a specially built cell orientating flow cytometer (OFCM). The design and operation of the OFCM are described. Measurements of the angular dependence of fluorescence from acriflavine stained rabbit sperm show that it is capable of orienting flat sperm with a tolerance of +-7/sup 0/. Differences in the angular dependence for the similarly shaped bull and rabbit sperm allow discrimination of these cells. We show that DNA staining with 4-6 diamidino-2-phenylindole (DAPI) or an ethidium bromide mithramycin combination allows resolution of the X and Y populations in mouse sperm. They have also been successful with sperm from the bull, ram, rabbit, and boar. Reliable results with human sperm are not obtained. The accuracy of the staining and measurement techniques are verified by the correct determination of the relative content of these two populations in sperm from normal mice and those with the Cattanach (7 to X) translocation. Among the potential uses of these techniques are measurement of DNA content errors induced in sperm due to mutagen exposure, and assessment of the fractions of X and Y sperm in semen that may have one population artifically enriched.

• Research Organization: Lawrence Livermore National Lab., CA
• OSTI ID: 6253737
• Journal Information: Cytometry (Baltimore); (United States), Journal Name: Cytometry (Baltimore); (United States) Vol. 3:1; ISSN CYTOD
• Country of Publication: United States
• Language: English

Related Subjects

550300* -- Cytology
59 BASIC BIOLOGICAL SCIENCES
ACCURACY
ACRIDINES
ACRIFLAVINE
AMINES
ANIMALS
AZINES
CATTLE
CELL CONSTITUENTS
CELL FLOW SYSTEMS
CELL NUCLEI
CHROMOSOMES
DATA
DNA
DOMESTIC ANIMALS
EXPERIMENTAL DATA
FLAVINES
FLUORESCENCE
GAMETES
GERM CELLS
HETEROCHROMOSOMES
HETEROCYCLIC COMPOUNDS
HISTOLOGICAL TECHNIQUES
INFORMATION
LUMINESCENCE
MAMMALS
MEASURING METHODS
MICE
NUCLEIC ACIDS
NUMERICAL DATA
ORGANIC COMPOUNDS
ORGANIC NITROGEN COMPOUNDS
ORIENTATION
PYRIDINES
RABBITS
RODENTS
RUMINANTS
SPERMATOZOA
STAINS
USES
VERTEBRATES
X CHROMOSOME
Y CHROMOSOME