PRACTICUM TOPIC

LIQUID/CHILLED SEMEN
PROCESSING

TEAM OF LECTURERS
DIVISION OF REPRODUCTION & OBSTETRICS
DEPARTMENT of VETERINARY CLINIC, REPRODUCTION AND PATHOLOGY
IPB UNIVERSITY
FACULTY OF VETERINARY MEDICINE STUDENT
 LEARNING OUTCOME

After learning the practicum topic of 9,

students are capable to
• calculate the semen extension for a
liquid/ chilled semen
• diluting of liquid semen for artificial
insemination purposes
 PRACTICUM PROCEDURES
1) Preparation of diluents
No. 1 - 3 have been
2) Semen storage implemented in
the previous
practicum
3) Evaluation of fresh semen

4) Implementation of dilutions TOPIK 9

5) Storage

6) Utilization:
Liquid semen for AI
Check the quality of liquid semen regularly
Daily Observations
 IMPLEMENTATION OF LIQUID SEMEN DILUTION

1. UNDERSTANDING THE CHARACTERISTICS OF ANIMAL’S SEMEN

2. HAVE DONE AN EVALUATION AND SEMEN STORAGE
3. CALCULATION OF THE DILUTION/EXTENSION PROCESS:
semen processing calculation formula is :

TV =Vo X Sperm concentration X motilitas (Formula 1)

AI Dose
TV = EV + Vo ..................................(Formula 2)
therefore it can be calculated EV = TV – Vo
Where:
TV = Total volume; Vo= semen volume ; EV = volume of extender
 CHILLED SEMEN
Characteristics of CHILLED semen

Criterian Liquid Semen

AI Dose
Insemination dose and volume
Bull 5-15 juta (0.5-1.0 mL)
Buck/ram 50-150 (0.05-0.20 mL)
Type of diluent/extender Fresh milk; Skim; Coconut water
Egg yolk (EY) & NaSitrat EY
Storage Refrigerator (temp. 3-5o C)
Life spent 3-4 days
 DILUTION OF BULL SEMEN
• Will process bull semen with volume 8 mL; Concentration of 800 x106
sperm / mL and 80% sperm motility
• Artificial Insemination Dose :
AI dose: 5-15 x106 sperm / 0.5 - 1.0 mL
means the number of cells / vol units in the straw:
Number of cells: 5-15 x106 sperm
Volume of semen: 0.5-1.0 mL
AI dose : 10 x106 sperm / 0.5 mL
Calculation :
Total Volume of Dilution = Vo X Sperm concentration X sperm motility
AI Dose

= 8 ml X 800 X 106 / mL X 80%

10X106/0.5 mL
= 8 X 800 X 106 X 80%X 0.5 ml
10X106
= 256 mL
 DILUTION OF BULL SEMEN
Calculation :
Total volume of dilution = Vo X Concentration of sperm X sperm motility
AI Dose
= 8 ml X 800 X 106 / mL X 80%
10X106/0.5 mL
= 8 X 800 X 106 X 80%X 0.5 ml
10X106
= 256 mL

Extender Volume = TV – Vo = 256 mL – 8 mL = 248 mL

This semen dilution can be used for AI:

TV/ AI dose volume = 256 ml / 0.5 mL = 512 heads of cows/ AI Dose

Naturally in seminal plasma consist of seminal plasmin (natural
antimicrobe)

livestock semen due to urinary tract along with the reproductive

tract (Urogenitalis), the bacteria naturally exist in semen

Adding extender to the semen

Seminal plasmin unable to protect
sperm anymore

Therefore we have to add an-antibiotics

To prevent contamination due to bacterial growth in
diluted semen.
To suppress bacterial growth
Antibiotics used: Penicillin and Streptomycin

Antibiotic Dose Penisilin 500-1000 IU/ mL chilled semen

Streptomycin 0.5-1.0 mg/ mL chilled semen

Antibiotic

3 x 106 Internasional Unit (IU) Streptomycin 1 mg

ANTIBIOTIC PREPARATION
Antibiotic stock preparation
(dilute with sterile Aquabidest, put mark / label on
antibiotic vial: date of dissolution, concentration (amount
/ mL of solution)

Penicillin 3,000,000 I.U dissolve into 15 mL

200,000 I.U / mL (Penicillin antibiotic stock)

Streptomycin 1 g (1000 mg) dissolve into 5 mL

200 mg / mL (Stock of antibiotic strep)

For semen that has been diluted with a total volume : 256 mL
is required :
Penicillin: 200X103 IU / mL stock,
The penicillin dose used is 1000 IU / mL of semen solution.

256 ml X 1000 IU/mL = 256 X 103 IU, from AB stock need to

be taken:
256 X 103 IU X 1mL = 1.28 mL
200X103 IU/mL
Streptomycin : Stok 200 mg/mL , the dose of streptomycin
used is 1 mg / mL of semen solution.

256 ml X 1 mg/mL = 256 mg, from AB stock need to

be taken :
256 mg X 1mL = 1.28 mL
200 mg
 Semen technique mixing
extender+ antibiotic (mix well) + finally the semen

Semen 8 mL Penicillin Streptomycin

extender 248 ML 1.28 ml 1.28 ml
A
B C
C add to B, homogenize
Than A
 PRACTICUM TOPIC 9 - LMS Report
Preparation of diluents

Semen storage
SIMU-
LATION
Fresh semen evaluation

Determination of dilution
CALCULATE
Storage

Daily Observations TO BE INFORM

TO BE KNOWN:
LIQUID SEMEN STORAGE WITH 4 DILUENT

DILUENT (HAVE BEEN PREPARED)

1. PURE/FRESH MILK
2. SKIM MILK
3. EGG YOLK NaSITRATE
4. EGG YOLK COCONUT WATER

LIQUID SEMEN THAT HAS BEEN MADE, STORED IN A TUBE LOCATED IN BEJANA
CONTAIN WATER (WATER JACKET) IN THE REFRIGERATOR WITH TEMPERATURE 3-5 ° C

SPERMATOZOA MOTILITY OBSERVATION IS CONDUCTED EVERY

24 HOURS UNTIL 0% MOTILITY

NOT IMPLEMENTED IN THE PRACTICUM TOPIC 9 - ONLINE / LMS

 REPORT
• INDIVIDUAL - FULL - use your own sentence, not plagiarism
• DISCUSSION ANSWERING THE QUESTIONS BELOW :

NOTED THAT:
Semen volume 0.8 ml
Concentration 2400 x 106/ml
Sperm Motility 75%

1. What kind of semen characteristics are the data

above?
2. Explain the calculation steps to make semen
processing until become chilled semen, and how
much Extender volume is needed?
3. How many females can be inseminated?