The Canine Brain and Tissue Bank has been collecting tissue samples from family dogs since 2017.
Through this period we have obtained brains and other samples from 118 dogs from various breeds, 5 wolves and 19 jackals.
List of animals, from which RNAlater stabilized samples (<4 hours post mortem) are available
| Tissue type / region | Storage method | Number of animals | Breeds | Other species | Remark |
| Whole brain (right hemisphere) | Simply frozen at -80°C as a whole (old sampling procedure) | 60 | Beagle(10), Bichon havanaise (2); Border Collie(3); Boxer(1);Chihuanua(1); Dachshund(1);Dobermann(1); German Shepherd(1); Golden retriever(2); Gordon Setter(1); Vizsla(2); Labrador retriever(5);Miniature poodle (2); Puli (1); Small münsterlander(1); Yorkshire terrier(2); Mongrel (variable,20) | Quality of samples may be questionable in cases where pieces have already been cut out from the brain tissue. We keep track of freeze-thawing cycles. | |
| Whole brain (right hemisphere) | Sliced and parts frozen separately at -80°C (current method) | 4 | Beagle (1); Borzoi(1); Whippet(1); Mongrel(1) | ||
| Whole brain (or left hemisphere) | Whole hemisphere fixed in 4% formaldehyde, stored at 4°C | 135 | Beagle(13); Bichon bolognese(1); Bichon frise(1); Bichon havanaise(3); Bobtail(1); Border Collie(3); Borzoi(1); Boxer(5); Caucasian Shepherd Dog(1); Chihuanua(1); ; Dachshund(1); Dobermann(1); English Bulldog(2); English Cocker Spaniel (1); German Shepherd(2); Golden retriever(3); Gordon Setter(1); Labrador retriever(6); Leonberger (1); Miniature poodle (2); Mudi(1); Puli (2); Rottweiler(1); Small Münsterlander(1); Staffordshire Terrier (1); Vizsla(2); Westie (2); Whippet(1); Yorkshire terrier(5); Mongrel (variable, ~30) | Canis aureus (19) and Canis lupus (1)*, please ask for details | |
| Brain / frontal cortex | Whole brain simply frozen at -80°C (1th phase sampling) | 60 | Beagle(10), Bichon havanaise (2); Border Collie(3); Boxer(1);Chihuanua(1); Dachshund(1);Dobermann(1); German Shepherd(1); Golden retriever(2); Gordon Setter(1); Vizsla(2); Labrador retriever(5);Miniature poodle (2); Puli (1); Small münsterlander(1); Yorkshire terrier(2); Mongrel (variable,20) | ||
| Brain / frontal cortex | Sliced and frozen at -80°C (current method) | 4 | Beagle (1); Borzoi(1); Whippet(1); Mongrel(1) | ||
| Brain / frontal cortex | RNA later stabilized tissue piece | 64 | Beagle(11), Bichon havanaise (2); Border Collie(3); Borzoi(1); Boxer(1);Chihuanua(1); Dachshund(1);Dobermann(1); German Shepherd(1); Golden retriever(2); Gordon Setter(1); Vizsla(2); Labrador retriever(5);Miniature poodle (2); Puli (1); Small münsterlander(1); Whippet(1); Yorkshire terrier(2); Mongrel (variable,21) female | Canis lupus (4), please ask for details | Some samples have already been used up for research, please ask for details. |
| Brain / frontal cortex | Fixed in 4 % formaldehyde as whole brain hemisphere | 135 | Beagle(13); Bichon bolognese(1); Bichon frise(1); Bichon havanaise(3); Bobtail(1); Border Collie(3); Borzoi(1); Boxer(5); Caucasian Shepherd Dog(1); Chihuanua(1); ; Dachshund(1); Dobermann(1); English Bulldog(2); English Cocker Spaniel (1); German Shepherd(2); Golden retriever(3); Gordon Setter(1); Labrador retriever(6); Leonberger (1); Miniature poodle (2); Mudi(1); Puli (2); Rottweiler(1); Small Münsterlander(1); Staffordshire Terrier (1); Vizsla(2); Westie (2); Whippet(1); Yorkshire terrier(5); Mongrel (variable, ~30) | Some samples have already been used up for research, please ask for details. | |
| Brain / cerebellum | Whole brain simply frozen at -80°C (1th phase sampling) | 60 | Beagle(10), Bichon havanaise (2); Border Collie(3); Boxer(1);Chihuanua(1); Dachshund(1);Dobermann(1); German Shepherd(1); Golden retriever(2); Gordon Setter(1); Vizsla(2); Labrador retriever(5);Miniature poodle (2); Puli (1); Small münsterlander(1); Yorkshire terrier(2); Mongrel (variable,20) | ||
| Brain / cerebellum | Sliced and frozen at -80°C (current method) | 4 | Beagle (1); Borzoi(1); Whippet(1); Mongrel(1) | ||
| Brain / cerebellum | RNA later stabilized tissue piece | 64 | Beagle(11), Bichon havanaise (2); Border Collie(3); Borzoi(1); Boxer(1);Chihuanua(1); Dachshund(1);Dobermann(1); German Shepherd(1); Golden retriever(2); Gordon Setter(1); Vizsla(2); Labrador retriever(5);Miniature poodle (2); Puli (1); Small münsterlander(1); Whippet(1); Yorkshire terrier(2); Mongrel (variable,21) female | Canis lupus (4), please ask for details | Some samples have already been used up for research, please ask for details. |
| Brain / cerebellum | Fixed in 4 % formaldehyde as whole brain hemisphere | 135 | Beagle(13); Bichon bolognese(1); Bichon frise(1); Bichon havanaise(3); Bobtail(1); Border Collie(3); Borzoi(1); Boxer(5); Caucasian Shepherd Dog(1); Chihuanua(1); ; Dachshund(1); Dobermann(1); English Bulldog(2); English Cocker Spaniel (1); German Shepherd(2); Golden retriever(3); Gordon Setter(1); Labrador retriever(6); Leonberger (1); Miniature poodle (2); Mudi(1); Puli (2); Rottweiler(1); Small Münsterlander(1); Staffordshire Terrier (1); Vizsla(2); Westie (2); Whippet(1); Yorkshire terrier(5); Mongrel (variable, ~30) | Some samples have already been used up for research, please ask for details. | |
| Brain / brain stem | Whole brain simply frozen at -80°C (1th phase sampling) | 60 | Beagle(10), Bichon havanaise (2); Border Collie(3); Boxer(1);Chihuanua(1); Dachshund(1);Dobermann(1); German Shepherd(1); Golden retriever(2); Gordon Setter(1); Vizsla(2); Labrador retriever(5);Miniature poodle (2); Puli (1); Small münsterlander(1); Yorkshire terrier(2); Mongrel (variable,20) | ||
| Brain / brain stem | Sliced and frozen at -80°C (current method) | 4 | Beagle (1); Borzoi(1); Whippet(1); Mongrel(1) | ||
| Brain / brain stem | RNA later stabilized tissue piece | 64 | Beagle(11), Bichon havanaise (2); Border Collie(3); Borzoi(1); Boxer(1);Chihuanua(1); Dachshund(1);Dobermann(1); German Shepherd(1); Golden retriever(2); Gordon Setter(1); Vizsla(2); Labrador retriever(5);Miniature poodle (2); Puli (1); Small münsterlander(1); Whippet(1); Yorkshire terrier(2); Mongrel (variable,21) | Canis lupus (4), please ask for details | Some samples have already been used up for research, please ask for details. |
| Brain / brain stem | Fixed in 4 % formaldehyde as whole brain hemisphere | 135 | Beagle(13); Bichon bolognese(1); Bichon frise(1); Bichon havanaise(3); Bobtail(1); Border Collie(3); Borzoi(1); Boxer(5); Caucasian Shepherd Dog(1); Chihuanua(1); ; Dachshund(1); Dobermann(1); English Bulldog(2); English Cocker Spaniel (1); German Shepherd(2); Golden retriever(3); Gordon Setter(1); Labrador retriever(6); Leonberger (1); Miniature poodle (2); Mudi(1); Puli (2); Rottweiler(1); Small Münsterlander(1); Staffordshire Terrier (1); Vizsla(2); Westie (2); Whippet(1); Yorkshire terrier(5); Mongrel (variable, ~30) | Some samples have already been used up for research, please ask for details. | |
| Skin from head | RNA later stabilized tissue piece | 64 | Beagle(11), Bichon havanaise (2); Border Collie(3); Borzoi(1); Boxer(1);Chihuanua(1); Dachshund(1);Dobermann(1); German Shepherd(1); Golden retriever(2); Gordon Setter(1); Vizsla(2); Labrador retriever(5);Miniature poodle (2); Puli (1); Small münsterlander(1); Whippet(1); Yorkshire terrier(2); Mongrel (variable,21) | ||
| Skin from nose | RNA later stabilized tissue piece | 64 | Beagle(11), Bichon havanaise (2); Border Collie(3); Borzoi(1); Boxer(1);Chihuanua(1); Dachshund(1);Dobermann(1); German Shepherd(1); Golden retriever(2); Gordon Setter(1); Vizsla(2); Labrador retriever(5);Miniature poodle (2); Puli (1); Small münsterlander(1); Whippet(1); Yorkshire terrier(2); Mongrel (variable,21) | ||
| Musculus temporalis | RNA later stabilized tissue piece | 64 | Beagle(11), Bichon havanaise (2); Border Collie(3); Borzoi(1); Boxer(1);Chihuanua(1); Dachshund(1);Dobermann(1); German Shepherd(1); Golden retriever(2); Gordon Setter(1); Vizsla(2); Labrador retriever(5);Miniature poodle (2); Puli (1); Small münsterlander(1); Whippet(1); Yorkshire terrier(2); Mongrel (variable,21) | Some samples have already been used up for research, please ask for details. | |
| Ggl. trigeminale | RNA later stabilized tissue piece | 64 | Beagle(11), Bichon havanaise (2); Border Collie(3); Borzoi(1); Boxer(1);Chihuanua(1); Dachshund(1);Dobermann(1); German Shepherd(1); Golden retriever(2); Gordon Setter(1); Vizsla(2); Labrador retriever(5);Miniature poodle (2); Puli (1); Small münsterlander(1); Whippet(1); Yorkshire terrier(2); Mongrel (variable,21) | ||
| Ggl. distale | RNA later stabilized tissue piece | 42 | Beagle (7); Border Collie (2); Bichon Havanaise (2); Boxer (1); Chihuahua (1); Dachshund(1); Dobermann(1); German Shepherd (1); Golden Retriever (2); Gordon Setter (1); Labrador Retriever (3); Miniature poodle (2); Small Münsterlander (1); Vizsla (2); Yorkshire Terrier (2); Mongrel (12) | Standard collection was discontinued from 2019. | |
| Ggl. Cervicale (C4 or C5*) | RNA later stabilized tissue piece | 42 | Beagle (7); Border Collie (2); Bichon Havanaise (2); Boxer (1); Chihuahua (1); Dachshund(1); Dobermann(1); German Shepherd (1); Golden Retriever (2); Gordon Setter (1); Labrador Retriever (3); Miniature poodle (2); Small Münsterlander (1); Vizsla (2); Yorkshire Terrier (2); Mongrel (12) | Standard collection was discontinued from 2019. | |
| Gl. thyreoidea | RNA later stabilized tissue piece | 42 | Beagle (7); Border Collie (2); Bichon Havanaise (2); Boxer (1); Chihuahua (1); Dachshund(1); Dobermann(1); German Shepherd (1); Golden Retriever (2); Gordon Setter (1); Labrador Retriever (3); Miniature poodle (2); Small Münsterlander (1); Vizsla (2); Yorkshire Terrier (2); Mongrel (12) | Standard collection was discontinued from 2019. | |
| Ln. retropharyngeus medialis | RNA later stabilized tissue piece | 42 | Beagle (7); Border Collie (2); Bichon Havanaise (2); Boxer (1); Chihuahua (1); Dachshund(1); Dobermann(1); German Shepherd (1); Golden Retriever (2); Gordon Setter (1); Labrador Retriever (3); Miniature poodle (2); Small Münsterlander (1); Vizsla (2); Yorkshire Terrier (2); Mongrel (12) | Standard collection was discontinued from 2019. | |
| Liver | RNA later stabilized tissue piece | 4 | Standard collection started from 2020. | ||
| Tumors | RNA later stabilized tissue piece | Samples collected on ocassion, ask for more details! |
Donation system
Donating the bodies of euthanized pet dogs is a voluntary action based on the concordant decision of the dogs’ owners and the veterinarians who would perform the euthanasia. Both the owners and veterinarians have to fill a Donation Document, which confirms the ownership of the dog and that it had valid vaccination against rabies, before the CBTB can accept the actual donation .
Sampling
In all cases when the CBTB gets informed about an expected donation, the transportation of the cadavers are arranged before the euthanasia takes place in order to minimalise the elapsed time between the death of the animal and the fixation of tissue samples. Therefore, we have been able to obtain molecular grade (stabilized in RNAlater, Thermo Fisher) tissue samples within four hours post mortem from 61 animals so far. These samples are being stored at -80 °C for long term. After taking tissue pieces from each target area (see below) and putting them in RNAlater, we process the whole brain: the right hemispheres get stored directly at -80 °C and the left hemispheres get fixed in 10% formaldehyde solution, and stored at +4 °C.
In other cases, where the post mortem delay exceeds this limit, only the brains of the animals are collected and get stored primarily for histological purposes.
Available samples
Cases with <4 hours post mortem delay
Tissue samples, which have been fixed in RNAlater within 4 hours post mortem, are generally available from several areas.
The locations from where we have collected RNAlater fixed tissue pieces from all animals (cases with <4 hour post mortem time) are the following: nose skin; skin from head; temporal muscle; trigeminal ganglion (whole ganglion); prefrontal cortex (gyrus compositus anterior); cerebellum; brain stem.
We also collect and store liquor cerebrospinalis, when it is obtainable (e.g. the animals were not extensively dehidrated, in which cases the liquor has a very limited accessible volume).
Buccal swabs and hair are also taken from all animals.
Furthermore, tissue pieces from some animals (<4 hours port mortem) from the following locations are also available: thyroid gland; retropharyngeal medial lymph node; liver and heart.
Cases with > 4 hours post mortem delay
We have formaldehyde fixed brains from several animals. We can provide information about these sample on demand.
Research and Perspectives
Our own research goals mainly focus on canine aging. We are currently investigating gene expression differences on the mRNA level, both by specific (targeting previously described genes) and high-throughput (RNA sequencing) approaches.
Through the first years of establishing the CBTB sampling and sample processing protocols we have monitored the quality of the samples by RT-qPCR, comparing the stability of house keeping genes (GAPDH, HMBS and HPRT1) between donated animals with different sampling times (within the 4 hours limit). As we haven’t detected correlation between RNA quality and and sampling time within this time frame, we plan to extend the time limit for collecting molecular grade samples. This will allow us to assign more donated animals for RNA based research goals.
Challenges and limitations
Our protocol which can allow efficient stabilization and subsequent purification of proteins for various research goals is still under development. As the RNAlater fixed samples are primarily intended for RNA analyses and there seems to be a demand for them, we would like to collect and store extra samples from the same animals, which will be primarily intended for proteomics investigations.