Fertility Testing

Frequencies of Defects among infertile couple
Ovulation Defects
Semen Abnormality -
Tubal defects
other disorders
Most Commonly used diagnostic steps for infertility
1. Semen analysis incld sperm antibody
2. Ultrasonographic scan of the pathway
3. Sperm function test
4. Testosterone level and other hormones
1. Endometrial biopsy
2. Serum progesterone estimation
3. Laparoscopy
4. Hysterosalpingogram
5. Post coital test
I. SEMEN Analysis
- failure to conceive after 6mos. of unprotected
- sexual abstinence of 3 days but not longer than 5 days
- longer abstinence = inc. semen vol. but reduced sperm
collect 2nd specimen in another 2 hours
- collect by masturbation
- evacuate bladder prior to ejaculation
- use pre-weighed sterile plastic container with screw top
- deliver specimen w/in an hour of collection
- if suspecting retrograde ejaculation
get post ejaculate urine
- collect and analyse 2 specimens at 2 to 3 wks interval
(+) marked difference, collect additional
Initial Macroscopic exam done after liquefaction
- liquefy in <20min at room temp.
failure to liquefy = inadequate prostatic
add proteolytic enzymes (bromelin, plasmin or chymotrypsin)
- thoroughly mix specimen before subsequent exam
and viscosity recorded.
Volume – 2 to 5 ml
Color – white or gray-white
yellowish hue = pyospermia
rust color = small bleedings in the seminal vesicle
3. pH – 7.2 to 7.8
8.0 or above = acute infection in the prostate, SV or
7.0 or lower = contamination with urine
= obstruction in the ED
= mainly prostatic fluid
Initial Microscopic Exam
1. Estimate of Sperm concentration Depth of 20um
4uL of sample
Total sperm ct = concentration x volume
(20 to 50 mil/mL)
(2 to 5 mL)
2. Motility = 50% or above
= done on warm stage
Grade 4 = rapid movement in a straight line with little
yaw and lateral movement
Grade 3 = slower
Grade 2 = slower (normal range)
Grade 1 = no forward progression
-motility of <50% = use viability stain of eosin with
nigrosin (red dye accumulation in the
head of non-motile sperm)
- samples with no visible sperm = centrifuge entire
= repeat analysis in another 4 to 6 months
3. Agglutination
- motile sperm stick to each other in a variety of orientation
- suggestive of immunologic cause of infertility
- record description of the type agglutination
Other Cells:
1. Round Cells
immature germ cells
(single or double highly condensed nucleus
with larger area of surrounding cytoplasm)
(smaller and inc. N/C ratio)
(+) peroxidase staining
2. Bacteria
3. Epithelial cells
-Negative sperm with low semen volume
fructose test
examine post ejaculate urine
Teratozoospermic index –average number of defects/spermatozoa
- acrosomal cap should be >1/3 of head surface
- cytoplasmic droplet of < half of the head size
- tail should be >45um long
II. Immunologic Assays
- antibody binding to head and tail is considered specific
for immunologic infertility.
- usually of the IgA or IgG and rarely IgM
- MAR test and immunobead assay
Latex bead
Human IgG
Increased risk of antibody
in men:
(a) vasectomy
(b) repeated infections
(c) obstruction of the ducts
(d) cryptochidism
(e) varicocele
(f) testicular biopsy
(g) trauma
(h) torsion
(I) cancer
(j) genetic predisposition
in women:
(a) intense mucosal inflammation of the genital tract
Microbiologic Assays
- E.coli can cause sperm agglutination
and immobilization
- >1000 cfu/ml
Biochemical Assays
Prostate gland – Zinc, Citric Acid, pH, & acid
phosphatase activity
Seminal Vesicle – fructose
Ejaculatory duct obstruction, Agenesis of Vas
defferens and SV - low volume, low pH,
no coagulation and no characteristic
semen odor
Epididymis – neutral alpha glucosidase
Fertilizing Function or Sperm Function Test
1. Sperm Penetration Assay (SPA)
- tests sperm fertilizing capacity
-acrosome rxn
-Speroolemma fusion
-sperm incorporation in the ooplasm
-decondensatin of sperm chromatin
2. Hemizona Assay
- uses unfertilized oocytes
- strict criteria morphology and IVF rates
3. Acrosin Assay
- trypsin-like serine proteinase
4. Hypo-osmotic swelling test (HOS)
- membrane integrity of sperm membrane
- correlates with SPA
5. Cervical Mucus Penetration Assay
- at midcycle
- inc . estrogen
- inc. water
- inc. Ca+2
- inc. pH (7 – 8.5)
Therapeutic Procedures
1. Sperm Wash
- IVF, GIFT, intrauterine insemination,
retrograde ejaculation, (+) sperm antibody
- separation within an hour of ejaculation
2. Semen Cryopreservation
- sperm banking
- before beginning cancer tx, before surgical
sterilization, life threatening situation, low
sperm count