Canine Brucella IFA Test

BACKGROUND:

Dog brucellosis, primarily caused by the bacterium Brucella canis, is a contagious disease affecting domestic dogs and wild canids.  The bacterium spreads through contact with infected animals or their bodily fluids, including mating, maternal transmission to puppies, and exposure to aborted fetal materials or vaginal discharges.  Infected dogs often exhibit reproductive issues such as infertility, failure to conceive, and abortion, with females typically aborting between 45 to 55 days of gestation. Males may suffer from epididymitis, testicular atrophy, and poor semen quality. General symptoms like lethargy, swollen lymph nodes, and fever can also occur, though they are less common.

Diagnosis of dog brucellosis involves serological tests to detect antibodies, culture tests to isolate the bacterium, and PCR to identify bacterial DNA. Treatment is challenging and often involves extended courses of antibiotics, though it may not fully eradicate the infection. Preventive measures include routine testing of breeding dogs, maintaining strict hygiene in kennels, and careful regulation of dog imports from regions with lower breeding standards. While the disease is zoonotic, transmission to humans is rare, typically resulting in symptoms such as fever and joint pain. Preventive practices and good hygiene can significantly reduce the risk of human infection.

TEST PRINCIPLE:

Dilution and Application:

• Dog sera are diluted in phosphate-buffered saline (PBS) with a pH of 7.2–7.4.
• The diluted sera are applied to slide wells.
• Incubation at 37°C allows for an antigen-antibody reaction if the sample is positive.

Washing:

• Non-bound nonspecific serum proteins are washed off with PBS.

Addition of Conjugate:

• Fluorescein-labeled FITC anti-dog IgG conjugate is added.
• The conjugate binds to the antigen-antibody complexes.

Incubation:

• Samples are incubated for 30 minutes.
• Unbound conjugate is washed off with PBS.

Preparation for Evaluation:

• Wells are covered with Mounting Medium.
• A cover slip is placed on top.

Evaluation:

• Samples are examined using a fluorescence microscope with a FITC filter system at 400× magnification.

MATERIALS REQUIRED:

Slides:

• 10 slides coated with Brucella canis antigens

Dropper Bottles:

• 1 dropper bottle with 3.0 ml FITC anti-dog IgG conjugate
• 1 dropper bottle with 0.5 ml Positive Control
• 1 dropper bottle with 0.5 ml Negative Control
• 1 dropper bottle with 3.0 ml Mounting Medium

Documentation:

• 1 instructions for use

MATERIALS REQUIRED BUT NOT PROVIDED:

Buffers and Solutions:

• PBS (phosphate-buffered saline) pH 7.2–7.4

Containers and Storage:

• Washbasin for PBS
• Test tubes for serum dilutions

Pipettes and Cover Slips:

• micro litre pipettes
• 24 x 50 mm cover slips

Microscope:

• Fluorescence microscope with filter system for FITC (fluorescein isothiocyanate)
  • Excitation wavelength: 465–495 nm
  • Barrier filter: 515–555 nm
  • 400× magnification

Incubation Equipment:

• 37°C incubator
• Humid chamber

SPECIMEN APPLICATIONS:

• Serum or plasma

STORAGE AND STABILITY:

Storage Temperature:

• Store the entire test kit at +2–8 °C.

Component Labels:

• Storage temperatures indicated on individual component labels apply if components are purchased separately (resulting in different expiry dates).

Shelf Life:

• The shelf life is 12 months from the date of manufacturing.

RECOMMENDATION OF USE:

Sample Correlation:

• Ensure test-kit components are correctly correlated to each specific dog sample.

Pipette Tips:

• Use a new pipette tip for each sample or dilution to avoid cross-contamination.

Conjugate Handling:

• Store the conjugate in the dark at 2–8 °C as it is photosensitive and heat-sensitive.
• The conjugate contains Evans-blue dye, which is potentially carcinogenic. Avoid ingestion and skin contact.

Infectious Material:

• Treat sample material and slides as potentially infectious. Dispose of them properly after the test procedure, along with used test-kit components.

TEST LIMITATIONS:

Seroprevalence:

• The cut-off value can vary based on the region and the origin of the sample (dependent on prevalence and the state of endemicity).
• It is recommended that each laboratory determines its individual cut-off.

Acute Infection:

• An acute infection (2- to 4-fold titre increase, known as “seroconversion”) can only be determined by comparing titres from a paired serum test (2 samples taken 2–3 weeks apart).

Test Result Interpretation:

• Always interpret test results in conjunction with anamnestic (patient history) and clinical data, as well as additional laboratory parameters.

Possible Cross Reactions:

• Cross reactions with sera positive for Brucella ovis, Brucella melitensis, and Brucella abortus are possible.

TEST PROCEDURE:

Preparation:

• Ensure all test-kit components (except the conjugate) and the sera to be tested are at room temperature.
  

Serum Dilution:

• Prepare appropriate dilutions (e.g., 1:40 and 1:80) with PBS for all sera to be tested.
• For previously positive sera, prepare serial two-fold dilutions in PBS to determine the endpoint titre (highest dilution that is still positive).

Application:

• Remove the slides from their foil pouch just before use and place them into the humid chamber.
• Apply 1 drop of the Positive and Negative Control on separate antigen wells on each slide.
• Pipette 20 μL of each serum dilution onto separate antigen wells, ensuring the wells are completely covered.

Incubation:

• Incubate the slides for 30 minutes at 37°C.

Washing:

• Gently tap off remaining serum dilutions from the slides.
• Shake the slides gently for 5 minutes in PBS, then repeat with fresh PBS for another 5 minutes.
• Briefly rinse the slides with distilled water, gently tap off remaining water, and dry the Teflon mask between wells if necessary (without letting the antigen wells dry out).
• When using a washing bottle, avoid directing the stream onto the antigen wells.
  

Conjugate Application:

• Place the slides back into the humid chamber.
• Immediately drop 1 drop of FITC anti-dog IgG conjugate onto each used well, ensuring the wells are completely covered.

Incubation:

• Incubate for 30 minutes at 37°C in the dark to protect the photosensitive conjugate.

Repeat Washing:

• Repeat the washing step as described in step 5.

Mounting:

• Add drops of Mounting Medium on the cover slips and place them carefully on the slides, removing any bubbles.

Evaluation:

• Evaluate the slides using a fluorescence microscope at 400× magnification, comparing each well to the fluorescence patterns of the Positive and Negative Controls.

Storage:

• Sealed slides can be stored at 2–8 °C in the dark for up to 7 days.

READING TEST RESULTS AND INTERPRETATION:

Microscope Requirements:

• Use a fluorescence microscope with a FITC filter system (excitation wavelength 465–495 nm, barrier filter 515–555 nm) and 400× magnification.

Reference Patterns:

• The fluorescence pattern (form, density, etc.) of the Negative and Positive Controls is used as the reference.
• Patterns of reactivity that differ from those seen in the Controls are considered non-specific and thus negative.

Positive Fluorescence Pattern (≥ 1:40):

• The coccobacilli exhibit clear, yellow-green fluorescence.
• Further dilution of positive samples is recommended to determine the endpoint titre (highest dilution that is still positive).

Cut-off Fluorescence Pattern / Recommended Cut-off (1:40):

• The coccobacilli show weak (1+) yellow-green fluorescence.

Negative Fluorescence Pattern (< 1:40):

• No clear fluorescence is visible.

Divergent Fluorescent Reaction:

• Reaction patterns different from those seen in the Positive Control are considered non-specific and thus negative.

DISCLAIMER OF LIABILITY:

• The entire risk associated with the performance of this product is assumed by the purchaser.
• The manufacturer is not liable for any indirect, special, or consequential damages resulting from the use of this product.

Canine Brucellosis FAQs

What is canine brucellosis?

• Canine brucellosis is an infectious disease caused by the bacterium Brucella canis. It primarily affects the reproductive organs of dogs, leading to infertility, abortions, and other reproductive issues. It can also cause other systemic symptoms and is zoonotic, meaning it can be transmitted to humans.

How is canine brucellosis transmitted?

• The disease is primarily spread through contact with bodily fluids from infected dogs, including blood, urine, semen, and vaginal discharges. It can also be transmitted through breeding, aborted fetuses, or contaminated food and water.

What are the symptoms of canine brucellosis?

• Common symptoms include reproductive issues such as infertility, spontaneous abortions, and stillbirths. Other signs can include lethargy, lameness, swollen lymph nodes, and chronic pain. Some dogs may show no symptoms but still carry the disease.

How is canine brucellosis diagnosed?

• Diagnosis is typically made through blood tests that detect antibodies against Brucella canis. Additional tests, such as bacterial culture and polymerase chain reaction (PCR), can be used to confirm the diagnosis.

Can canine brucellosis be treated?

• Treatment for canine brucellosis is challenging and often involves a combination of antibiotics over an extended period. Despite treatment, some dogs may remain carriers of the bacteria. Infected dogs are often recommended to be neutered or spayed to prevent the spread of the disease.

Is canine brucellosis contagious to humans?

• Yes, Brucella canis is zoonotic and can infect humans. People in close contact with infected dogs, such as breeders, veterinarians, and pet owners, are at higher risk. Human infection can cause flu-like symptoms, and in severe cases, more serious health issues.

How can the spread of canine brucellosis be prevented?

• Prevention includes regular screening and testing of breeding dogs, practicing good hygiene, and isolating infected animals. It is also important to avoid breeding dogs that have tested positive for Brucella canis.

What should I do if my dog is diagnosed with canine brucellosis?

• Consult your veterinarian for a comprehensive treatment plan. Follow their guidance on managing the infection, which may include isolation from other animals, medication, and potential lifestyle adjustments. Inform anyone who has had contact with your dog, including breeders and other pet owners.

Can canine brucellosis be prevented through vaccination?

• Currently, there is no vaccine available for Brucella canis. Prevention relies on screening, testing, and good hygiene practices to reduce the risk of infection.

Are certain breeds more susceptible to canine brucellosis?

• Canine brucellosis can affect any breed of dog. However, it is more commonly seen in breeding kennels and among dogs with close contact with other dogs, such as in shelters or dog shows. Regular screening in such environments is crucial.

Canine Brucella Antibody Rapid Test FAQs

What is the Canine Brucella Antibody Rapid Test?

• The Canine Brucella Antibody Rapid Test is a diagnostic tool used to detect antibodies against Brucella canis in dogs. It is a quick and reliable method to identify exposure to the bacteria, which can cause brucellosis in canines.

Why is it important to test for Brucella canis in dogs?

• Brucella canis is a bacterial infection that can lead to reproductive issues, lameness, and other health problems in dogs. It is also zoonotic, meaning it can be transmitted to humans. Early detection helps in managing and controlling the spread of the disease.

How is the Canine Brucella Antibody Rapid Test performed?

• The test is performed by taking a small blood sample from the dog, which is then applied to the test device. Results are typically available within 10-15 minutes, making it a rapid and convenient option for veterinarians.

What do the results of the test mean?

• A positive result indicates the presence of antibodies against Brucella canis, suggesting that the dog has been exposed to the bacteria. A negative result means no antibodies were detected, implying no exposure or an early stage of infection where antibodies are not yet detectable.

Can a dog test positive for Brucella canis but not show any symptoms?

• Yes, some dogs may be asymptomatic carriers of Brucella canis, meaning they can test positive for antibodies without showing clinical signs of the disease. This makes regular screening important, especially in breeding or multi-dog environments.

How reliable is the Canine Brucella Antibody Rapid Test?

• The test is highly reliable, with sensitivity and specificity values generally above 90%. However, confirmatory testing using additional methods like PCR or culture is recommended for positive cases.

Can the test differentiate between active and past infections?

• The Canine Brucella Antibody Rapid Test detects antibodies, which indicates exposure but cannot distinguish between active and past infections. Further testing and clinical evaluation are necessary to determine the infection status.

Is there any preparation required before testing?

• No special preparation is required for the dog before the test. However, it is important to follow the test kit instructions carefully to ensure accurate results.

What should I do if my dog tests positive for Brucella canis?

• If your dog tests positive, consult with your veterinarian for a confirmatory test and discuss the appropriate treatment and management plan. It's also important to take measures to prevent the spread of the infection to other animals and humans.

How often should dogs be tested for Brucella canis?

• The frequency of testing depends on the dog's risk factors. Breeding dogs, those in shelters, or in multi-dog households should be tested regularly, typically every 6-12 months. Consult with your veterinarian to determine an appropriate testing schedule for your dog.