Before you inseminate you must have your work bench well layed out, and all equipment sterile. Laboratory grade tissues and are very handy for cleaning up contamination.
Sterility is of the upmost importance, any contamination can give the queen and infection and she will surely die. Glass tips, tweezers, forceps etc are autoclaved (this can actually be done in a pressure cooker if no autoclave available).
You try to get yourself in the habit of moving always from left to right so every task is carried out methodically.
Drone Husbandry in the Lab:-
Once you have collected your mature drones in drone cages (wooden cage with queen exluders either side, about 100 per cage), they must be nurtured. Drones without workers get cold very fast, they become hard to evert as well as many simply will drop dead.
The best method is to make a drone flight box with a heated lamp. A box with an elastic sleeve inside is a cage containing workers with a candy food source. You simply release the drones inside and secure the sleeve. This box serves many purposes. Firstly as the drones were excluded in the colony they were unable to fly and defecate (which is what they like to do) In the cage they can do this, so the drones are very clean and this reduces a possible contamination source. Secondly they fly around then return to the cage surface containing nurse bees and are fed. This keeps them energetic at all times resulting in an easy eversion during the semen collection period.
Buffer solution:- This is a saline based solution containing an antibiotic. There are many different recepies for this. It is personal preference. The purpose of the solution is a backing solution inside the semen collection syringe and tip. You need it to prevent drying of the semen and to assist semen delivery by capillary action. Some people also collect a drop of this solution between drones to prevent drying from the other end. This is then expelled from the tip when collecting from the next drone.
Drone semen collection:-
Once you have mastered this you are well on your way. The process requires you to firstly achieve the partial eversion of the drone and then the full eversion using pressure.
The drone must be tense to achieve this properly. A good way to achieve this is by turning the drone upside down in one hand and grip it between the thumb and forefinger. Apply pressure to the abdomen and remove or crush the head on the drone
with your other hand. This helps the drone tense. By now you should have acheived partial eversion. Maintain pressure and gently roll your fingers downwards until the full eversion takes place. Many drones will pop fast, it is important to learn to control the speed of the full eversion. If the semen sac bends right over and touches the body of the drone, this is another potential contamination. These drones are simply discarded into your prepared rubbish bin. I EXPECT YOU ALL TO GIVE THIS A TRY, AFTER ALL WHEN A DRONE MATES IT'S WHOLE APPENDAGE IS TORN OFF AND THE SIMPLY BLEED TO DEATH. CRUSHING THE HEAD IS MORE HUMANE IN MY EYES.
Once you have the full eversion it's time to collect the semen. You must have your apparatus and prepared syringe under a microscope with sufficient power. You will see a ball of pinky white liquid on the surface of a white fluffy mucus layer. The idea is to skim the pink semen with the use of capillary action from the mucus layer. Mucus in the tip can be devestating, It clogs the tip and is a possible contaminent.
Firstly you wind the buffer solution up and down and then all the way out of the tip and make sure you have good capillary action. This is vital to make semen collection smooth. You retract this solution about 5mm back up the tip to maintain an airspace between the buffer and the semen. You make contact with the largest portion of the pink layer, wind foward slightly to make contact and then begin retracting removing semen until the majority is removed. A big tip, don't try to take all of it. you are destined to get mucus. A good mature drone contains approximately 1 microlitre of semen. 8 microlitres are used to inseminate carniolans and 10 microlitres for italians (it varys throughout the world).
The process is repeated untill you fill a glass tip. You wind the sperm down the tip to make contact with the next drone and then repeat capillary action. The glass tips I use contain enough sperm for inseminating 4 queens. I collect all semen first this may be multiple tips. The filled tips can be removed and remain perfectly viable for up to 48 hrs. However the fresher the better. Never attempt to do more queens than you can handle.
Drone quality is extremely variable. Good prepared drones like I've mentioned make the semen collection process smoother. Some drones just simply don't have any sperm. It can often take 15-50 drones per queen depending on you level of preparation.
This is like writing a novel, the next chapter is coming soon