Professional Documents
Culture Documents
Artificial Insemination:
Semen Processing and Quality Control
The objective of a boar stud is the efficient production
of adequate quantities of quality semen doses of
appropriate genetics, shelf- life and health. Achievement
of this objective demands practical expertise and a high
degree of attention paid to the details. Every process on the
boar stud must be described to ensure standardization of
excellent practice. A number of studs have no w achieved
ISO9002 certification. This involves a great deal of work, but
is a clear demonstration of commitment to quality assurance,
and provides a framework for ensuring maintenance of the
highest standards.
Ideally, the AI lab should be close to the semen collection area, and fitted with pass-
through windows so that semen may be transferred rapidly. On larger studs, where this is
not possible, semen may be delivered to the lab via a pneumatic tube delivery system.
The laboratory should be designed with labor efficiency and work flow in mind. While
providing adequate work surface and storage, the lab needs to be compact, to minimize
the distance between work stations, and also the amount of cleaning.
Consumable items
Semen collection, evaluation, processing and dispatch utilize large quantities of single-
use disposable items e.g. collection bags and/or cups, filters, gloves, plastic pipettes,
microscope slides, tubing, extender bags and/or vats, dispatch boxes and/or bags. These
must all be of high quality and adequate for the purpose, and are best purchased from a
reputable AI equipment manufacturer. Whilst it may be tempting to reduce costs by
purchasing a lower grade cheaper alternative, great care must be taken to ensure that
anything used will not damage the semen. You must check that each item is non-
spermicidal. You must also consider the quality of an item and weigh this against the
cost. Finally, you need a reliable source of consumables, and have a back-up plan in case
there is ever a problem with supply.
Semen evaluation
Every aspect of AI demands a high degree of quality control and consistency. A fixed
protocol must be followed in order to ensure semen of consistently high quality. Semen
should be examined immediately after collection. Throughout semen processing, every
effort should be made to avoid rapid changes in temperature. Remember that all
processing steps are a balance between speed and accuracy.
Sperm count
This is usually measured using some form of photometer calibrated for the purpose,
which is quick and easy to use. Two popular models are the MicroReaderT M (IMV) and
the SpermacueT M (Minitube). The major disadvantage of any photometric method of
estimating sperm count is that it measures optical density only, and cannot therefore
Motility estimation
This provides a measure of sperm viability. Motility may be evaluated for the raw
ejaculate, for partially and fully extended semen. It provides an estimation of the
proportion of sperm cells showing progressive forward motility. This parameter is
important because it has an impact on the subsequent fertility of the dose.
Prepare a microscope slide by warming it to approximately 950 F (350 C). Place one
standard-sized drop of semen onto the slide using a pipette, and place a warm cover slip
over the drop. Use a microscope fitted with a warm-stage at 950 F (350 C) and examine
the sperm cells at x200 or x400 magnification to estimate the level and vitality of sperm
movement.
Examining raw semen, it is possible only to give the quality of sperm motility a score
according to a simple scale (e.g. from 1-5, where 5 is excellent, 3 is acceptable for further
processing, less than 3 to be discarded).
Where partially or fully extended semen is examined, attempt to estimate the proportion
of sperm that are active, and the quality of that activity, rating it in terms of the
percentage of sperm showing progressive forward motility. Note that in extended semen
samples, it is easier to visualize individual sperm cells than in a raw ejaculate as the
density is lower. In this case, a minimum of 60% motile sperm is essential for good
levels of fertility with AI.
Some of the larger studs in the US are now adopting Computer Aided Semen Assessment
(CASA) systems such as IVOST M from Hamilton Thorn Research, or SpermVisionT M
from Minitube. These systems utilize computer technology to track individual sperm
cells, evaluating various parameters of their movement (e.g. direction, velocity, angle of
curvature between the head and the tail) and can provide a much more accurate, objective
and repeatable measure of both sperm count and motility. They represent a significant
investment initially, and also require a commitment to train staff properly, but yield
benefits in terms of accuracy in semen evaluation, reduction of sperm wastage and labor
saving in the long-term.
Some of the computerized systems for semen evaluation can also classify sperm
morphology.
Semen extension
Semen should be extended as soon as possible after collection. Most studs aim at
a sperm dose of 2.0-4.0 x 109 viable sperm in 75-100mls. extender. The number of doses
made from an ejaculate is calculated as follows:
For example:
No. sperm per ml semen: = 0.525 x 109
Volume of ejaculate = 120 mls
No. sperm required per dose: = 3.0 x 109
= 21
(No. sperm doses x Volume of each dose) – Total volume of the ejaculate
For example:.
No. semen doses: = 21
Volume of each dose: = 90 mls
Volume of ejaculate = 120 mls
= 1770mls
Semen Extenders
The functions of the semen extender are to expand the volume of the ejaculate, and to
maintain sperm cell viability. Extenders may be prepared from raw ingredients in the AI
laboratory, but this is not recommended. High precision weigh-scales are required in
order to do the job properly; doing the job yourself may compromise quality control, and
is not cost effective. Most AI studs purchase their semen extenders in powder form.
There is a wide range of semen extenders available commercially, which vary in
performance and price. They include BTS, MerckIIIT M, VitalT M, AndrohepT M, XcellT M,
MR-AT M, Androhep EnduraguardTM and SafeCellT M.
The choice of extender depends on the shelf- life required, and the price you are prepared
to pay. Whilst the formulation of many extenders is now either patented or confidential,
it is important that you have some information about this from both health and safety, and
quality assurance viewpoints. Also, it is essential that you are clear about the antibiotic
content of the extender you are using. If any problem should arise regarding the
bacteriology of the semen doses you may need to alter the antibiotic content of the
extender, and this is impossible without knowing what is added to start with.
Development of new diagnostic tests for the presence of virus in semen have been
developed. PCR (Polymerase Chain Reaction) technology is extremely sensitive, and can
identify presence of very low levels of virus in a sample. The PCR test for PRRS virus in
semen works well, and many studs now send semen samples for testing on a routine
(weekly) basis. The turnaround time for the test is rapid, with results being available
within hours of receipt at the lab. Where PRRS is an issue, it would be possible to hold
semen doses in storage for 24 hours, dispatching to farms only after a negative PRRS
PCR test result has been confirmed.
Water quality
A dose of semen is at least 90% water! The quality of water used for the preparation of
the semen extender is very important, but the choice of processes can be confusing. In
simple terms, the following water purification methods are appropriate:
Water may be purified in the AI lab, and this is common practice for larger studs. The
best method for your laboratory will depend upon the quality of the raw water at the point
of delivery. AI equipment manufacturers can provide guidance on the type of processing
plant best suited to your individual requirements. Where purified water is purchased,
make sure you are satisfied with the process employed, and the quality control adopted
by the supplier. Note that the quality of purified water deteriorates with time, and
according to the method of storage. When purchasing purified water, make sure that no
more than 1 month’s supply is on site at any one time, to avoid deterioration of water
quality.
Extender vats are available for larger-scale operations, with a capacity of 100+litres.
They are made of stainless steel, usually lined with a disposable plastic liner, and are
fitted with a mixer and a thermostatically-controlled heater. They can be installed onto a
weigh-cell to simplify filling with the required amount of purified water. Usually the vat
is filled the day before, and the heater set to switch on during the night, ha ving water at
the correct temperature in time for semen processing the following morning. Extender
powder is then added at the start of collections, allowing time to equilibrate before use.
Once semen has been dispensed into doses, it must be cooled to 59-640 F (15-180 C) prior
to dispatch. The cooling process should be gradual (1-2 hours from dispensing into bags
or tubes), to avoid damaging the sperm cells.
Equipment manufacturers
The equipment and consumable items mentioned in this text is available from:
IMV
11725 95th Avenue North
Maple Grove, MN 55369, USA
Phone: 763-488-1881, Fax: 763-488-1888
IMV
BP 81 61302 L’Aigle Cedex, France
Phone: +33 2 33 34 64 64, Fax: +33 2 33 34 11 98
Kubus SA
Calle E, s/n
Pol. Ind. C/E Europolis
28230 Las Rozas
Madrid, Spain
Phone: +34 1 636 0268, Fax: +34 1 637 5313
Minitube of America
P.O. Box 930187
Verona, WI 53593, USA
Phone: 800-646-4882, 608-845-1502, Fax: 608-845-1522
Minitub GmbH
Hauptstrasse 41
84184 Tiefenbach b. Landshut, Germany
Phone: +49 8709 9229 0, Fax: +49 8709 9229 39