Cleanliness                    Return to part 1 of this article
By B. W. Pickett, Ph.D.
Professor Emeritus, Colorado State University

Part 2:
When semen is collected with an AV, there is always concern that the semen will not truly represent the physiological condition of the stallion. In short, was the collector and/or his/her equipment or handling of the semen responsible for poor semen quality or were the spermatozoa deficient due to the stallion. Unfortunately, the relationship between motility and fertility of the sperm is not always reliable. Occasionally, motility of the sperm will be highly satisfactory immediately after the semen is collected, but may not be fertile, or fertility will decrease rapidly even though motility remains excellent. Therefore, the damage to the sperm may not be immediately evident, but shows up later, which is called latent damage. Dust, dirt and residues from soap and/or other cleaning agents, semen, smegma, lubricants, etc., that were not removed after previous collections can result in poor quality semen, particularly when the spermatozoa is from a stallion with marginal fertility.

Ideally, the combination liner and cone, filter and collection bottle, those objects that come into contact with semen, should be clean and sterile. Unfortunately, this is rarely possible under practical farm conditions. Consequently, about the best we can strive toward is that they are clean, which is much more important than their being sterile.

Cleaning Latex Artificial Vagina Liners
Although it never occurs to many of the personnel in the breeding barn, but most rubber is porous, particularly the type of latex used for collection of semen. Thus, when the liners are new chemical residues may remain on the latex and in the pores. Therefore, latex liners should be cleaned (conditioned) very, very thoroughly before they are used the first time. The following is the procedure we have used for years, and certainly appears satisfactory.

1.) Turn the combination liner and cone (liner) inside out, i.e., so that the side that comes into contact with semen is to the outside.

2.) Place them in a washing machine (not the washing machine in the barn). Set the water on warm/warm and the dial on heavy load. Use the longest cycle with an extra rinse, if available.

3.) Put the liners through five complete cycles using a detergent that contains no dyes or fragrances.

4.) Using the same settings as 2, repeat for five cycles, but do not use a detergent. This is in essence tap water rinses.

5.) Place the liners lengthwise in a plastic container that can be covered. Add 3 to 5 gallons of distilled water, depending upon the number of liners being "conditioned."

6.) The liners should be completely submerged. This can be done by placing heavy objects, such as bricks in individual zip-lock bags, on top of the liners.

7.) After the liners have remained submerged in distilled or deionized water at least 24 hours, they should be rinsed.

8.) Rinse at least two additional times in distilled or deionized water.

9.) Hang them in a dust-free cabinet to dry. Building and maintaining a dust-free cabinet in a laboratory with direct access to a room where semen is collected, or some other part of a barn is not an easy task. Ideally, the room where liners are washed and stored, should be separate from where the semen is brought from the collection shed
Note: In the event you feel that this procedure is too troublesome. Preconditioned liners can be purchased from Animal Reproduction Systems.

The following method should be used to clean liners after they have been used.

1.) Remove the liner from the artificial vagina. This can be done by placing the AV upside down in a sink, after the collection bottle has been removed.
Note: The water in the AV should have been removed immediately after collecting semen from the stallion.
Put water into the open end until approximately 2/3 full. When the casing is raised quickly, the pressure of the water will invert the liner so that the "dirty" side is outside. Then remove from the cuff on the posterior end.

2.) Clean the liner with cold, running water until the rubber is "squeaky" clean of smegma, semen, lubricant, hair, etc. Rubber is porous, therefore only cold water should be used during the first cleaning step. Hot water is likely to "set" the proteinaceous material onto the latex rubber.

3.) The liners are then placed in a container of Nolvasan® solution (1:49 Nolvasan® to water) and rinsed. The liners can remain in the Nolvasan® solution for up to 24 hrs, but they should be completely submerged, or drying at the junction of the air and solution will result in deposition of residue, which is exceedingly difficult to remove. However, if this occurs and is not removed, it will contaminate the semen.

4.) Remove liners from the Nolvasan® solution and place them in a sink. A three compartment sink with coved corners is almost ideal. The first compartment should contain hot water and Terg-A-Zime®, which is a biodegradable enzyme detergent, highly soluble in water. In this compartment, the liners are scrubbed with a brush.

5.) The second compartment contains warm tap water. The liners are rinsed in this compartment four times, changing the water each time. The water should be sufficiently warm to dissolve all the detergent, but not hot enough to make the liners "sticky."

6.) In the third compartment is cool distilled or deionized water. The liners are rinsed four times, changing the water between rinses.

7.) The liners are then hung in a dust-free cabinet to dry.

8.) When thoroughly dry, put on a pair of wrist-length plastic gloves and turn the liner outside in, so they will be ready to use.
Note: Do not use alcohol in an attempt to "sterilize" the liners. It is ineffective, because it collects water and is no longer at the concentration necessary to kill bacteria. Further, most alcohols that are readily available will leave a residue on the rubber.

Remember, all soaps and detergents, as well as water, are spermicidal. Therefore, all material that comes into contact with the semen should be free of chemicals and residues; thus, the reason for numerous rinses and drying thoroughly. We recognize that many of our readers will think that all the rinsing that we have recommended is excessive. However, it has been recommended that all glassware and equipment used in embryo transfer be rinsed 12 times in tap water and 12 times in distilled, deionized water.

When the breeding season is over, all rubber ware should be thoroughly cleaned, dried, placed into zip-lock bags and stored in a deep-freeze [(-20ºC) (-4ºF)]. They will stay clean under these conditions, and, most important, the rubber will not age. As rubber ages, it becomes more porous (cracked), and is more difficult to clean.

On many farms today, in addition to handling semen for immediate use and/or shipping, semen is also being frozen. Ideally, these areas should be separated.

To reduce microbes, dust and any chemical contaminants, it is recommended that laboratory work surfaces be washed regularly as follows: soapy water, then a clear water rinse, followed by a mild disinfectant that is relatively nontoxic. Surfaces must be allowed to dry thoroughly before use.

No effort has been made to describe methods for cleaning glassware or plasticware. Unfortunately, it is as difficult to clean and keep clean. Fortunately, there are numerous companies that manufacture and/or sell disposable material that is generally sterile and inexpensive. Consequently, we recommend that breeding farms use disposable material for everything, except latex liners for collection of semen. However, it should be noted that plastic, disposable liners are available and should be used for collection of semen whenever the stallion will readily ejaculate quickly and completely when they are used. Unfortunately, not all stallions will ejaculate completely without numerous intromissions into a plastic liner.

This Concludes Part 2:
Believe me there is no easy road to success when it comes to getting mares pregnant, and as you well know that road is paved with failures that took short cuts..

Go to part 3 of this article

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