AUBF_Finals_Lec: Semen Analysis

Parameters of Semen Analysis

• Appearance

• Vol

• Viscosity

• Ph

• Sperm conc.

• Sperm count

• Motility

• Morphology

• Vitality

Purpose of Semen Analysis

• Determine infertility

• Success of vasectomy or vasovasostomy

• Forensic analysis

Composition of Semen

• Spermatozoa

• Seminal Fluid

• Prostate Fluid

• Bulbourethral glands

This structure is respoinsible for spermatogenesis

Seminiferous tubules

Structure: for sperm maturation

Epididymis

Structure: propel sperm to ejaculatory ducts

Ductus deferens

Provide nutrients (ex. Fructose) for sperm and fluid

Seminal vesicles

StructureL Provides enzymes and proteins for coag and liquefactio

Prostate gland

add alkaline mucus to neutralize prostatic acid and vaginal acidity

Bulbourethral glands

Sexual abstinence

• at least 2 days but not more than 7 days

Storage of seme spx

Kept at room temo, must be processed within 1 hr of collection

What to do during delay in analsysis

37 C temp

Ways for spx. collection of Semen

Masturbation

Coitus interruptus

Coitus

Assisted ejaculation

directing semen into a clean sample cup (no lubricant)

Masturbation

withdrawing before ejaculating into a clean sample cup

Coitus interruptus

using a condom

Coitus

electro-ejaculation used in paraplegics

•Assisted ejaculation

Appearance of semen

Gray-white color, translucent

ODor

:musty odor (bleach-like)

How to liquify semen if it didnt after 2 hrs?

Add equal am’t of Dulbecco’s phosphate buffered saline (DPBS) or proteolytic enzymes (chymotrypsin/bromelain)

>2cm threads means?

highly viscous semen

normal pH semen

7.2-8.0

Dilution used for semen analysis (semen conc. and count)

1:20 dilution

Both sides of the hemocytometer are loaded and allowed to settle for__ - __ minutes.

3-5 mins.

Diluent used for sperm conc. and count

• Sodium bicarbonate and formalin / saline and distilled water

Purpose of diluent for Sperm conc and count

Immobilize the sperm

What kind of sperms are counted for sperm concentration and sperm counted

Matured sperms

Formula for Sperm Concentration

___ sperm counted x 1,000,000 = __ sperm/mL

Sperm count

Sperm/mL x (specimen volume aspirated) = ___ sperm/ejaculate

Sperm motility Preparation and conditions needed upon doing this procedure?

1. Liquefied semen

2. 10 uL under a cover slip, settle for 1 min

3. Observe in 20 HPF

Alternative sperm motility grading criteria

PM - Progressive motility

NO - Nonprogressive motility

IM - Immotility

Sperm motility Grading (by WHO)

abnormality in the head part will affect

ovum penetration

abnormal neckpice, midpeice, and tail will affect the sperm’s

the sperm’s motility

Preparation for Sperm morphologyg

• Thin smear

• 10 uL of semen

• Staining with Wright’s, Giemsa, Shorr, or Papanicolaou stain.

How many sperm should be evaluated in sperm morphology

At least 200 sperm should be evaluated, and the percentage of abnormal sperm is reported

Stains used in Sperm morphology

• Wright’s stain

• Giemsa Stain

• Shorr

• Papanicolaou stain

Conditions under sperm vitality

• assessed within 1 hr of ejaculation

• Mix teh specimen with an eosin-nigrosine stain

What are the causes of Low frcutose levels in seminal fluid fructose?

• abnormalities in seminal vesicle

• bilateral congenital absence of vas deferens,

• obstruction of the ejaculatory duct,

• partial retrograde ejaculation,

• androgen deficiency.

Test used to measure Seminal Fluid Fructose?

Resorcinol Test (Seliwanoff’s Test)

Positive color resulr for Seliwanoff’s test

Orange color

Normal level of Seminal Fluid

> 13 umol/ejaculate

Stain used for Sperm vitality

Eosin-nigrosine stain

Automated Semen Analysis Systems

• These are automated machines where:

• Objective determination of both sperm velocity and trajectory, including sperm concentration and morphology are included in the analysis

Additional testing for SA

Seminal Fluid Fructose

Antisperm antibodies

What is the appearance of (+) in Antisperm antibodies

There is clotting/Agglutination of sperm cells in microscopic exam

Test/s used to determine (+) in Antisperm antibodies

• Mixed Agglutination Reaction

• Immunobead test

• Detection of IgG

•Motile sperm incubated with IgG antihuman globulin (AHG) and a suspension of latex particles or Coomb’s cells

• + agglutination/clumps

Mixed Agglutination Reaction (MAR)

• Detects presence if IgG, IgM and IgA antibodies

•Sperm is mixed with polyacrylamide beads coated with anti-IgG, anti-IgM or anti IgA

• Bead attached to sperm at particular presence of beads on less than 50% of the sperm is considered normal

Tip:

Anti- = Antibody

Immunobead Test

Explain teh principle of Immunobead test

• Sperm mixed with Polyacrylamide beads

What kind of beads is used in immunobead test?

• Polyacrylamide beads

Normal percentage in immunobead test?

Less than 50%