Preserving the Specimen

Preserving the specimen is tricky and complicated process and should only be undertaken if you are confident in the process.  Anyone keen on learning the essential basics can contact one of the listed museums and a short demonstration of the fixation and specimen setting process will be conducted, as well as supplies of some basic field preservatives, DNA tissue solution, proper tags, etc.

Fixation involves the preservation and stiffening of tissues so that they do not decompose and so that the specimen remains in an easy-to-study posture. Preservation involves the storage of the specimen in a medium that allows for its safe study for many years.

Basic equipment needed is the following:

• A field note book or a reliable field data app on a phone or tablet
• Good quality specimens
• Surgical gloves to protect your hands from the chemicals
• Newspaper to work on
• Plastic fixing tray – a large, shallow plastic container is ideal with a polystyrene layer
• Syringes with needles (large volume syringes with large gauge needles for bigger specimens, as well as smaller syringes and finer needles for slighter body parts and smaller specimens) 
• Forceps, scalpels and surgical scissors for collecting tissue
• 70% ethanol for reptiles and amphibians
• 10% formalin ( 1 part 40% Formaldehyde + 9 parts water)
• Archival marker pens
• Tag paper, fine crochet thread and a needle for attaching tags to the specimens

Warnings: Formalin is known to cause cancer and it is important to minimize exposure to these chemicals when using them. Always work in an open space and wear gloves. The use of syringes with needles is another hazard associated with this protocol. Syringes are used to inject specimens with formalin (a carcinogen). Handle syringes carefully. Feel free to place the specimen being fixed in a tray to inject it instead of holding it in your other hand to minimize the risk of sticking yourself with the needle. If stuck by a needle, rinse with cold water for 15 minutes and wash area with soap and water. The final hazard associated with fixation and preservation process is the specimens themselves. Since these are dead animals, they may carry zoonoses and parasites. To minimize the hazard of being exposed to these pathogens, always handle specimens with gloves.

Prior to preserving the specimen, first collect the tissue sample. It should be emphasised that all instruments (scalpel, scissors, forceps) used for collecting tissue samples must be both thoroughly cleaned and sterilised with 70% alcohol (ethanol) before and after taking each sample, and that neither the working surfaces of the instruments nor the tissue being collected must be touched by the worker’s hands. To avoid contamination, putting labels inside the specimen container, while theoretically desirable as a safeguard against loss of data, carries a high risk of contamination and is therefore not recommended. The volume of fixative should be large, at least 20 times the volume of the specimen. The samples must be protected from light at all times and well labelled.

Fixing specimens:

• Fixation of specimens involves injection of formalin into the specimen, so should be done with caution and not in confined spaces
• Specimens should be fresh or thawed (if they were frozen).
• Place specimen in fixing tray, ventral side up.
• Use syringe to inject formalin into the body cavity, limbs and tail base.
  • Use a fine needle so as not to damage specimen.
  • Inject in two places in body cavity – in the abdomen and near the heart.
  • Inject enough formalin to mostly fill, but not over fill the cavities – if the body starts to distend, stop injecting.
  • Larger specimens can also be injected superficial in the limbs and tail, but volumes injected should be small, as there is no cavity in these body parts.
  • Snakes should be injected in multiple places along the body (5-8, depending on size).
  • Tortoises and terrapins should be injected near where the limbs enter the shell.
• Use the needle to prick through the skin of the limbs and tail to allow formalin to enter. One or two pricks per appendage is sufficient. Extrude (force out) the hemipenes if possible using formalin. Fine cotton can be used to tie at the base of the hemipenes to keep shape.
• The fixing tray should have a cloth that is soaked with formalin in the bottom.
• Position the specimen in the desired position whilst it is still supple.
  • Lizards & geckos: straight body; elbows, knees and ankles at 90 degrees, tail looped to one side.
  • Snakes: coiled into a spiral with the head inside, spiral can be ovular to ensure that specimen fits in jar.
  • Frogs: straight body, elbows at 90 degrees, knees and ankles <90 degrees.
  • Tortoises and terrapins: limbs and neck stretched out of shell, as possible, mouth wedged open with a paper wad.
• Tie a label with a field number onto the specimen with cotton thread. Record this in your field notebook or app.
  • Lizards, geckos and frogs: tie label on the limb or waste. Tortoises and terrapins, tie label to a hind limb, the bend of the knee is best.
  • Snakes: tie behind the head or at midbody, making sure the label cannot slip off
• Cover specimen with a cloth soaked with formalin and close tray. Let reptile specimen fix for 5 – 7 days and amphibiansfor 24-48 hours.
• Rinse specimen to remove formalin and place specimen in a basin of cold water and let it soak for 1-2 hours.
• Place specimen in used alcohol as a specimen that is freshly fixed and rinsed contains a lot of water, which will dilute the alcohol in a jar. Leave specimens in the used ethanol for 1 to 8 weeks.
• Place specimen in its permanent jar with new alcohol – always remember: 70% ethanol for reptiles and amphibians.
• Hand in the specimen to your preferred museum together with all the relevant documentation and the DNA tissue!

Note the position of the labels on each group. Frogs and geckos/lizards is optional between the two places often with smaller specimens labelled around the waste and larger specimens on the leg.

Permits

Check with your local museum whether their permit covers them receiving such specimens or allows for field collaborators to collect dead specimens on their behalf. Please note, that most provinces require permits, even for the collecting and transporting dead specimens. Museums or their officials will not legally place their institutions at risk if the collector fails to adhere to the relevant conservation legislations.  It should also be noted that if the specimen data is used in a research publication, the journal often requires proof that the specimen was legally collected (i.e. a permit valid for the period and locality from where the specimen was obtained). Failure to take this into consideration may result in the loss of valuable information to the scientific committee.

List of museums and institutions for specimen donations.

• Ditsong National Museum of Natural History, Pretoria, Gauteng.
  Contact Lauretta Mahlangu – mahlangu@ditsong.org.za

• National Museum, Bloemfontien, Free State
  Contact Mike Bates – herp@nasmus.co.za

• McGregor Museum, Kimberley, Northern Cape
  Contact Beryl Wilson – berylwa@museumsnc.co.za

• Port Elizabeth Museum, Port Elizabeth, Eastern Cape
  Contact Werner Conradie – wernerconradie@gmail.com

• Durban Natural Science Museum, Durban, KwaZulu-Natal
  Contact Leigh Richards – Leigh.Richards@durban.gov.za

• KwaZulu-Natal Museum, Pietermaritzburg, KwaZulu-Natal
  Contact John Midgley – Jmidgley@nmsa.org.za

• University of the Western Cape, Cape Town, Western Cape
  Contact Bryan Maritz – bmaritz@uwc.ac.za

• Rhodes University, Grahamstown, Eastern Cape
  Contact Shelley Edwards – S.Edwards@ru.ac.za

• The Venom Pit Snake Park, East London, Eastern Cape
  Contact Juan Marillier – thevenompit@gmail.com