SUBMISSION FORM for Tissue Collection Environmental Conservation Division, Northwest Fisheries Science Center, NMFS/NOAA, 2725 Montlake Blvd East, Seattle, WA 98112

Background Information: Organization submitting samples:______, Individual collecting samples:______

Animal identification/tag number and/or name:______

Species: Scientific name: Common name: ______

Dates of Stranding:______, of Death:______, of Tissue collection:______

Location of stranding:______

Name or location of the pod/colony the animal was from (if known or applicable):______

Age (in years, if known, or use neonate, pup/calf, juvenile, adult):______

Size: Length (snout to tail tip/fluke notch)______, Weight (estimate)______or (actual)______Axillary girth (half girth if large cetacean)______

Sex (if adult female please note reproductive status): Male____, Female____, Pregnant?____, Lactating?____, Neither___

Body condition (please check one): Emaciated_____, Fair_____, Good_____, Robust______Blubber thickness (measure over the xyphoid cartilage, excluding the skin)______, Oil leaching from blubber?___

Feeding status: Entire digestive tract empty______, Stomach empty but intestines contain some material______, Recent meal (please list type and amounts of stomach contents if ingested in the wild)______

Cause of death (if known):

Tissue Preservation: Wrap each tissue sample in a rinsed Teflon sheet and then place each bag in a separate labeled Whirl-pak® bag. Seal and place on ice. Freeze as soon as possible.

Tissues Requested: Please check which samples were taken and indicate the location taken from. _____1. Blubber - Full thickness (from skin to muscle) taken from over the xyphoid cartilage, if possible, 100-200 grams.

Location, if different than indicated: 2. Liver - 100 grams. Location sample taken from: _____3. Kidney - 100 grams. Location sample taken from:

Additional History (including behavioral observations, treatment records, and necropsy results, if available, any abnormalities found, and any other information available about the animal). Please use the back of this page and/or attach additional sheets and records. Thank You.

% , " Marine Mammal Tissue Sampling Protocol

Tissue Sampling for Chemical Contaminant Analyses

Supplies for sampling will include: — 12" x 13" acetone rinsed Teflon sheets — 18 oz. Whirl-pak® bags (4.5" x 8.5") — "Rite-in-the-Rain" tags for inside Whirl-pak® bags — rubber bands — ballpoint and marking pens — submission forms on "Rite-in-the-Rain" paper

Sampling Protocol • Priority for collection of samples is: blubber, liver, kidney (see attached Marine Mammal Sampling Sheet/Data Record for description of how and from where to collect tissue samples). • See attached Marine Mammal Tissue Sampling Protocol Addendum for additional information on sampling procedure. • For tissue collection, use a stainless steel knife and rinse the knife between necropsies of each animal. • Wrap individual tissue samples in Teflon sheet, secure with rubber band, and then put into labeled Whirl-pak® bags. Label the bags with: Animal ID Number______Species______Tissue Type______Date Collected______

• Insert labeled (as above) "Rite-in-the-Rain" tags into each Whirl-pak® bag (this is in case the writing comes off the Whirl-pak® bag). Remove as much of the air as possible from the bag before it is sealed. Place samples on ice. As soon as possible, freeze at lowest temperature available. • Please provide copy of full necropsy report (see attached Marine Mammal Sampling Sheet/Data Record).

Shipment of Samples • Ship frozen samples, early in the week, with ~5 lbs dry ice, FedEx overnight to: Karen Tilbury, NWFSC, ECD, 2725 Montlake Blvd. E., Seattle, WA 98112-2097. Call Karen (206-860-3338) the day the samples are shipped with the invoice number for tracking, if necessary.

% , " Marine Mammal Tissue Contaminant Analyses Environmental Conservation Division, Northwest Fisheries Science Center, National Marine Fisheries Service, National Oceanic and Atmospheric Administration, 2725 Montlake Blvd. East, Seattle, WA 98112-2097 Phone: 206-860-3330 FAX: 206-860-3335

MARINE MAMMAL TISSUE SAMPLING PROTOCOL

Tissue collection procedures - It is important to use standardized sampling procedures so that, even when there are low levels of contaminants present, the differences may be attributed to biological processes and contaminant exposure and not to variation in the collection process. The following procedures are essential to prevent cross-contamination of tissues within an animal and ensure uniformity of samples among animals.

1. Collect tissues only from animals that have been dead less than 24 hours. Even within 24 hours after death, the autolytic changes may become severe enough to confound efforts to interpret the contaminant data. This is why accurate information about the time between death and tissue collection is important.

2. Collect liver, blubber, and kidney samples from the locations specified on the submission form, if possible. This reduces variation caused by possible composition differences within tissues of the same animal. It also provides us with uniform samples and information from all participating organizations which can be directly compared based on the demographics of the animals. Sample size: blubber; 100 - 200 g (10 g minimum which is the full thickness of blubber and about the area of a credit card), liver; 100 g (size of a small human fist), kidney; 100 g (size of a small human fist).

3. Rinse all instruments with isopropyl alcohol before each tissue is sampled. This will minimize cross-contamination of tissues. Particular care needs to be taken after cutting through the skin (because of possible environmental contamination) and blubber (because of the high lipid content). When collecting blubber, remove the skin to prevent contamination, but include the full thickness of blubber from skin to muscle. Ideally, separate instruments would be used and scalpel blades changed after skin and blubber are cut.

4. Treat sample containers provided as if they are sterile. The teflon sheets have been rinsed with acetone, and then dried in a dust free environment before shipment to your facility. This is to reduce the possibility of external contamination before and during the sampling procedure by any of the compounds we are studying.

5. Keep samples as cold as possible after collection. Some of the organic contaminants are volatile or are degraded by compounds released during cell death, particularly in the liver and kidney. To decrease changes in contaminant levels due to these processes, keep the samples on ice following the necropsy and place in a freezer as soon as possible.

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