Semen Analysis

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Semen Analysis

• Clinical Pathology

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Semen Collection

• Semen is often collected into an artificial
  vagina, usually while a teaser bitch is present.
• An artificial vagina may be made of latex or
  disposable plastic.

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• May use electoejaculation in which a probe is
  attached to the pelvic nerves to stimulate
  ejaculation.

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Semen (3 fractions)

• 1st- mainly prostatic fluid, few sperm
• Released during the period of vigorous thrusting
• 2nd- Sperm rich portion of ejaculate
• 3rd- Mainly prostatic fluid, few sperm, majority
  of total volume of ejaculate.
• Buck, bulls, tom, ram you should collect all
  three
• Stallions, dogs, boars- collect 2nd and 3rd
  portions seperately

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Semen Handling Techniques

• Avoid marked temperature changes
• Avoid exposure to water, disinfectants
• Use clean, dry, warm equipment (37 C or 98 F)
• Slides, coverslips, pipettes, stains.
• Process soon after collection

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Semen Evaluation

• Color and consistency (normal is milky and
  moderately viscous)
• Volume
• Wave motion/sperm motility
• Spermatozoa concentration
• Morphology
• Ratio livedead
• Presence of foreign cells/material

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Volume of Ejaculate

• Measured in volumetric flask
• Method of collection affects volume
• Electroejaculation- volume is larger
• Teasing with a female-volume is larger
• Species variation
• Dogs 10-40 ml
• Stallion 65 ml
• Tom 0.5 ml
• Volume does not necessarily correlate with
  fertility

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Sperm Motility

• Motility correlates with fertility
• Improper handling can affect motility
• Evaluate immediately after collection
• Place a drop of semen on a warm slide,
  immediately cover with coverslip
• Dilute with warm saline if high concentration of
  sperm

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Classes of Sperm Motility

• Can be classified as good, very good, fair, or
  poor.
• Normal sperm should have greater than 70
  motility
• Examine under 100 x, may need to dilute
  concentrated samples
• Poor is when there is less than 40 motility

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Wave Motion

• Under low power 40x, look for swirling
• Progressive motility- sperm are moving around all
  over slide
• Non-progressive motility- sperm are only swimming
  in a similar pattern

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Sperm Concentration

• Most important characteristic
• Dilute a portion 1100 with saline or red cell
  Unopette.
• Using a hematocytometer, count total of sperm in
  the central grid
• Multiply the number by 2 million
• Boars/Stallions 150 M/ml
• Bulls 1200 M/ml
• Dogs 300 M/ml
• Cats 1700 M/ml

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LiveDead Sperm Ratio

• Place 1 drop eosin/nigrosin stain (make sure it
  is warm) and mix gently with a drop of semen on a
  warm slide.
• After several seconds, smear like blood smear.
• Live sperm resist staining-appear white against a
  blue-black background
• Dead sperm take up the eosin and stain pinkish
  red
• Examine and observe 200 cells

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Sperm Morphology

• Can examine on eosin/nigrosin stained smear
• Other stains india ink, HE, Wrights
• Observe 100-500 cells
• Record of abnormal cells
• Divide problems into head, neck, midpiece, and
  tail problems.
• Primary abnormalities occur during sperm
  production.
• Secondary occur from storage in the epididymis
  until the smear is made

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Double headed Sperm
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Misshapen Head
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Elongated Head
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Pear shaped head and bent midpiece
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Proximal Droplet
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Distal Droplet
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Detached Head
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Bent Tail or Midpiece
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Coiled Tail
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• http//www.vivo.colostate.edu/hbooks/pathphys/repr
  od/semeneval/conc.html