Sample Collection Protocols
SAMPLE COLLECTION PROTOCOLS
PURPOSE:
To measure the levels of Adrenocorticotropic Hormone (ACTH) circulating in the blood. Is useful in the differential diagnosis of pituitary Cushing’s disease, Addison’s disease, autonomous ACTH producing pituitary tumors (e.g. Nelson’s syndrome), hypopituitarism with ACTH deficiency and entopic ACTH syndrome.
PROCEDURE:
The determination of Endogenous ACTH MUST BE PERFORMED ON EDTA PLASMA. Do not use heparinized plasma samples. A minimum of 1.0 ml of EDTA plasma is required to perform the assay.
1. The animal should have been off medication for 1 week and fasted for at least 12 hours.
2. Collect blood into a chilled plastic syringe and Immediately transfer blood into a siliconized lavender top tube. These are special lavender top tubes and not regular lavender top tubes. Collection and storage into non-siliconized tubes will invalidate the test as ACTH binds to glass.
*Alternately, samples can be collected directly into a siliconized red top tube that has had EDTA added (3.75 mg/2.5 ml whole blood).
3. Separate plasma by placing in centrifuge for 3-5 minutes as soon as possible. Plasma must then be transfered into another siliconized red top tube using a plastic transfer pipet.
4. Freeze separated plasma sample immediately and transfer to lab frozen. Inform courier that you have a sample that must remain frozen.
Samples that are being shipped should always be shipped with ice packs. Dry Ice should be used if available.
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PRE-COLLECTION NOTES:
When considering this test, it is important that you use ACTH Gel or Cortrosyn (Organon) on a regular basis. Interpretation can be difficult when both are used interchangeably. Collection timing is very important as ACTH Gel and Cortrosyn are absorbed at different rates.
COLLECTION WITH ACTH GEL:
Canine - Collect sample 2 hours post-injection
Feline - Collect sample 1 hour post-injection
COLLECTION WITH CORTROSYN:
Canine - Collect sample 1 hour post-injection (0.25 mg IM)
Feline - Collect sample 30 minutes and 1 hour post-injection (1/2 vial)
PROCEDURE:
1. Animal should be fasted for at least 12 hours
2. Collect PRE sample into a red top tube or serum separator tube and label tube immediately, then refrigerate.
3. Administer either ACTH Gel or Cortrosyn as directed
4. Collect POST sample at appropriate time and label tube.
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PURPOSE:
To access the liver’s ability to metabolize bile acids after feeding. This is useful in evaluation hepatic dysfunction.
PROCEDURE:
1. Animal should be fasted for at least 12 hours.
2. Collect the PRE sample into a serum separator tube. Allow clot to form and centrifuge and refrigerate.
3. Feed the animal a fatty meal
4. Collect the POST sample into a serum separator tube. Allow clot to form and centrifuge and refrigerate.
*Make sure tubes are clearly labeled as PRE and POST feeding.
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PROCEDURE:
1. Animal should be fasted for at least 12 hours.
2. Collect PRE-Dex (High/Low) sample into a red top tube or serum separator and label tube immediately, then refrigerate.
3. Administer Dexamethazone (Low Dose- 0.01 mg/kg IV/High Dose - 1.0 mg/kg IV)
4. Draw POST 4 Hour and POST 8 Hour samples and label, then refrigerate.
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PURPOSE:
To evaluate an animal’s response to insulin therapy.
PROCEDURE:
1. Collect PRE sample into a gray top tube prior to administering insulin.
2. Collect additional samples at 2 hour intervals after administration of insulin.
*Make sure tubes are clearly labeled with the times samples were drawn.
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PROCEDURE:
1. Collect sample directly into a blue top tube. Allow tube to fill completely.
2. Inspect sample for clots. If clots are present, redraw sample.
3. Separate plasma from cells by placing in a centrifuge for 3-5 minutes or letting sit in a refrigerator for at least 3 hours.
4. Transfer plasma into a clean plastic tube or red top tube. Care must be taken when transferring so as not to transfer any cell as they will rupture when the sample is frozen and cause hemolysis.
5. Freeze sample immediately and inform courier that you have a sample that is frozen.
*If shipping sample, make sure you have adequate insulation and ice packs included.
Conditions that will invalidate the test:
Samples that are hemolyzed or if clots are present.
Do not test bitches in heat, pregnant, or lactating.
Dogs that are sick or have been on medication or vaccinated within the past 14 days.
Use of certain drugs that impair hemostasis.
Significant stress
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Contact lab for kit and full instructions 1 week prior to testing. Special collecting materials will be provided as needed.
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