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Sample Collection Protocol

Dual Path Platform (DPP) test evaluation in Mule and Sika Species

The United States Department of Agriculture (USDA), Animal and Plant Health Inspection Service

(APHIS), Veterinary Services (VS) is evaluating the effectiveness of the Dual Path Platform (DPP) as an 

official primary test and secondary test for bovine tuberculosis (TB) program use in captive and free 

ranging mule deer (Odocoileus hemionus) and sika deer (Cervus nippon). 

This guidance document represents the Agency’s position on this topic. It does not create or confer any 

rights for or on any person and does not bind the U.S. Department of Agriculture (USDA) or the public. VS 

may make this information available to the public. While this document provides guidance for users outside 

VS, VS employees may not deviate from the directions provided herein without appropriate justification 

and supervisory concurrence

Sample collection procedures for Mule and Sika DPP Evaluation

A. Test administration, ordering, and payment:

1) Designated accredited veterinarians(DAV’s)may perform the DPP sample collection for Mule 

and Sika deer

2) The National Veterinary Services Laboratory (NVSL) can provide serum submission kits if 

needed; call 515-337-6200 or email NVSL_userfee@aphis.usda.gov. The serum submission kit 

is not required to complete testing. Each kit contains shipping instructions, an ice pack, VS 

Forms 10-4 (Specimen Submission Form) and 10-4A (Continuation Sheet), and a prepaid 

shipping label to the NVSL. DAVs may also submit serum samples using their own shipping 

box, which should include ice or gel packs as well as the laboratory submission forms. The 

Office of Management and Budget approved VS Forms 10-4 and 10-4A for submitting all 

diagnostic samples to the NVSL. (See the VS Forms 10-4 and10-4A at 

http://www.aphis.usda.gov/library/forms/pdf/VS_Form10_4.pdf.)

3) Before drawing blood samples, the accredited veterinarian will:

a. Contact the VS Assistant Director (AD) in the animals’ State of location to verify that the 

State’s current animal health regulations allow use of the DPP serological tests for bovine 

TB in Mule or Sika deer. 

b. Contact the State Animal Health Official (SAHO) if testing is for interstate movement. 

Verify the receiving State will accept the DPP serologic test results for entry. 

B. Sample collection, processing, and shipping for DPP testing at NVSL:

1) Sample collection:

a. Collect blood samples in a 10-ml red top (clot) tube or 10 ml serum separation tube.

b. Collect 10 ml of whole blood to obtain 2 ml of serum to submit for testing.

c. DO NOT place freshly collected tubes directly on ice.

2) Sample processing and handling:

a. Allow the blood to clot at room temperature.

b. Once clotted, remove the serum from the clot and place it into a new, clean tube. If 

necessary, centrifuge the clotted blood sample to obtain at least 2 mL of serum. The 

United States

Department of

Agriculture

Animal and Plant

Health Inspection

Service

Veterinary Services
serum must contain minimal hemolysis. Place the serum tubes in a refrigerator until 

shipped to the NVSL. Do not freeze the serum.

c. Number and label submitted tubes with official identification so laboratory personnel can 

easily correlate the samples to the animal’s official ID as listed on the submission forms. 

This helps laboratory personnel verify receiving all the samples and correlate the results 

to individual animals. 

d. Organize the serum tubes into a tube box in the same order you listed the animals on the 

submission form or attached list. This allows the receiving laboratory personnel to 

quickly verify and test the submitted samples. If you need additional sample submission 

kits from the NVSL, obtain them before collecting the blood samples.

e. If also testing for brucellosis, submit a separate red top tube of blood to an approved 

brucellosis testing laboratory. The NVSL does not perform routine brucellosis 

surveillance testing.

3) Sample submission forms and TB test Record:

a. Fill out the VS Form 10-4 and VS Form 10-4A accurately and completely. (See the VS 

Forms 10-4 and10-4A http://www.aphis.usda.gov/library/forms/pdf/VS_Form10_4.pdf.).

1. Include official identification (and all other ID), species of cervid tested (Mule, 

Sika), age, and gender for each animal. 

2. You do not have to write animal identification on the submission form as long you 

attach a list with all of the above information. If preferred, attach to the VS Form 10-

4 a spreadsheet or printout of the animals with official ID corresponding to their 

blood tube numbers or attach a copy of the completed (all above information) VS 

Form VS 6-22 (Tuberculosis Test Record) with the corresponding blood tube 

numbers.

b. Submit the original VS Forms 10-4 and 10-4A to the NVSL as part of the sample 

submission described in Section B(4) of this Guidance.

c. Fill out the VS Form VS 6-22 (Tuberculosis Test Record) accurately and completely, 

identifying the cervid species tested (Mule or Sika), ID, age, and gender.

d. Submit the VS Form 6-22 to the State Veterinarian or VS AD in the State within 5 

business days of sample collection.

4) Sample submission to NVSL:

a. Pack submitted sample tubes with frozen gel packs. Do not put the tubes directly on the 

frozen packs in the container used to ship the samples to the NVSL. Refrigerate all serum 

samples until shipping. Do not freeze the serum.

b. Ship the samples along with the completed form VS 10-4/10-4A to the NVSL.

c. Ship serum samples using overnight or 2-day delivery. The NVSL recommends shipping 

the samples as quickly as possible (ideally within 24 to 48 hours), but no more than 72 

hours after collection. Do not ship the samples so they will arrive at NVSL on the 

weekend or a holiday. VS prefers that you use a shipper such as Federal Express or 

United Parcel Service if not using prepaid NVSL shipping containers, as they deliver 

directly to the NVSL facility. The U.S. Postal Service does not deliver directly to the 

NVSL.

d. The submitting veterinarian is responsible for payment of DPP tests conducted at NVSL. 

Three options are available for payment: call the business office 515-337-6200 and set up 
an account, provide a credit card number on the submission form, or include a check 

made out to NVSL with the samples and submission form. Diagnostic test rates can be 

found at 

https://www.aphis.usda.gov/animal_health/lab_info_services/downloads/AmesDiagnostic

TestingCatalog.pdf VS does not charge for the secondary DPP test after 30 days on 

animals that have primary positive test results.

e. Ship the serum samples to the NVSL at the following address:

National Veterinary Services Laboratories

1920 Dayton Avenue

Ames, Iowa 50010

5) Result interpretation and classification of animals:

a. The NVSL will test serum samples using the DPP following test kit instructions and 

internal NVSL standard operating procedures. If the NVSL does not have enough DPP 

test kits to perform the testing, the NVSL will freeze serum samples and notify the 

submitter of the delay. 

b. The NVSL will report an optical density value below 200 a negative DPP test result. The 

NVSL will report an optical density value equal to or above 200 a non-negative DPP test 

result.

c. The NVSL will report DPP primary test results to the submitting accredited veterinarian, 

the respective State animal health officials, the respective State VS AD, and the Cervid 

Health Commodity Team. 

Please direct any inquiries to: 

National Cervid Tuberculosis Disease Specialist 

USDA APHIS Veterinary Services 

Sheep, Goat, Cervid, and Equine Health Center – Cervid Health Program 

vs.sprs.cervid.health@aphis.usda.gov