FERTILITY TESTING

Frequencies of Defects among infertile couple

Ovulation Defects - 30%

Semen Abnormality - 22%

Tubal defects - 17%

other disorders - 12%

Endometriosis - 5%

Most Commonly used diagnostic steps for infertility

Men:1. Semen analysis incld sperm antibody2. Ultrasonographic scan of the pathway3. Sperm function test 4. Testosterone level and other hormones

Women:1. Endometrial biopsy2. Serum progesterone estimation3. Laparoscopy4. Hysterosalpingogram5. Post coital test

I. SEMEN Analysis- failure to conceive after 6mos. of unprotected

intercourse- sexual abstinence of 3 days but not longer than 5 days- longer abstinence = inc. semen vol. but reduced sperm

motility

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 specimens

Initial Macroscopic exam done after liquefaction

- liquefy in <20min at room temp.failure to liquefy = inadequate prostatic

secretion

add proteolytic enzymes (bromelin, plasmin or chymotrypsin)

- thoroughly mix specimen before subsequent exam and viscosity recorded.

Macroscopic

• Volume – 2 to 5 ml

• Color – white or gray-whiteyellowish hue = pyospermiarust color = small bleedings in the seminal vesicle

3. pH – 7.2 to 7.88.0 or above = acute infection in the prostate, SV or

epididymis7.0 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 sampleHemocytometer

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 = slowerGrade 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 sample

= 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 PMN(single or double highly condensed nucleus (smaller and inc. N/C ratio)with larger area of surrounding cytoplasm) (+) peroxidase staining

2. Bacteria3. 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

C/S

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

-capacitation-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