A Complete Guide to Quantitative D-Dimer Measurement Technique

Definition of D-Dimer

D-Dimer is generated as a specific soluble degradation product during fibrinolysis. A D-dimer test looks for D-dimer in blood. A D-dimer test is used to check for a blood clot. A blood clot is a mass formed when platelets, proteins, and cells in the blood clump together. When a person gets an injury, the body forms a blood clot to stop the bleeding. Blood clotting is an important process that stops one from losing too much blood when they are injured.

Significance of D-Dimer

Elevated D-Dimer levels are found in cases of:

Deep vein thrombosis (DVT), Here, a blood clot that builds up in a vein deep in the body. They normally affect the lower legs, but they can also happen in other parts of the body. DIC may be caused by inflammation, infection, or cancer.
Pulmonary embolism (PE), Here, a blockage forms in a lung artery. It happens when a blood clot in another part of the body breaks loose and travels to to the lungs. DVT clots are a common cause of PE.

Other circumstances may also lead to high D-Dimer levels such as:

Old age,
Pregnancy,
Cancer,
Liver disease
Infection.

The FIA (Fluorescence immunoassay) system has a higher sensitivity when compared with latex agglutination tests thus giving an improved aid in the diagnosis for the exclusion of venous thromboembolism (VTE).

Principle of D-Dimer test using FIA (Fluorescence immunoassay)

The test uses capture antibody for D-Dimer immobilized on a nitrocellulose strip. The detector antibody in conjugate clumps to the antigen in sample, forming antigen-antibody fluorescence complex. This complex migrates through the nitrocellulose strip by capillary action.

When the complex meets the line of the corresponding immobilized antibody on the test line, the fluorescence signal is produced from Immuno-complex. The greater the concentration of the antigen in sample forms the more antigen-antibody complex and leads to stronger intensity of fluorescence signal. The signal is interpreted and the result is displayed on analyzer in terms of concentration.

Requirements (Quanti Card Test kit)

D-Dimer Quanti Card Test kit has the following components to perform the assay:

D-Dimer Quanti Card (1 Test)
Assay Buffer
D-Dimer Conjugate (dried)
Clamp & Rod
iQuant Analyzer
Micro-pipette & Microtips
Stop Watch

Safety Warnings

The use of disposable gloves and proper bio-hazardous clothing is STRONGLY RECOMMENDED while running the test. In case is a cut or wound in hand, DO NOT PERFORM THE TEST.
Do not smoke, drink or eat in areas where specimens or kit reagents are being operated.
Tests are for in vitro diagnostic use only and should be run by competent personnel only.
Do not pipette by mouth.
Consult a physician immediately in case of accident or contact with eyes, in the event that contaminated material are ingested or come in contact with skin puncture or wounds.
Spills should be decontaminated promptly with Sodium Hypochlorite or any other suitable disinfectant.
D-Dimer conjugate and Assay Buffer contains Sodium Azide as a preservative. If these materials are to be disposed off through a sink or other common plumbing systems, flush with generous amounts of water to prevent accumulation of potentially explosive compounds. In addition, consult the manual guideline “Safety Management No. CDC- 22”, Decontamination of Laboratory Sink Drains to remove Azide salts
Follow standard bio-safety guidelines for handling & disposal of potentially infective material.
Use only Plasma samples collected using Sodium citrate as anti-coagulant.
All materials used in the assay and samples should be disposed off in accordance with established safety procedures. Wash hands thoroughly with soap or any suitable detergent, after the use of the kit.

In order to obtain reproducible results, the following instructions must be followed:

Use disposable gloves while handling potentially infectious samples and performing the
assay Wash hands thoroughly afterwards
Do not use the kit beyond the expiry date.
Do not mix reagents from different batches.
Do not open the foil pouch of the cards and D-Dimer Conjugate vial (Dried) until it attains room temperature (20-30°C).
Do not expose the D-Dimer Conjugate vial (Dried) to light for more than 8 hours.
Always reseal the unused D-Dimer Conjugate vial along with desiccant using Clamp and Rod provided and store at 2-8°C. Do not re-use the test device.
Use separate pipette tips for each sample in order to avoid cross-contamination of samples which could cause erroneous results.
Follow the given test procedure and storage instructions strictly to get accurate result

Sample collection and storage

Only plasma samples should be used with this test. It is recommended to use fresh plasma specimen for testing.
If the plasma specimen cannot be tested immediately, it should be refrigerated at 2-8°C for testing within 24 hrs. For storage more than 24hrs, freeze the specimen at -20°C or below.
Repeated freezing and thawing of the specimen should be avoided. Thawing of frozen sample should be done one time only, just prior to proceeding for test.
Specimen containing precipitates or particulate matter may yield inconsistent test results. Such specimens must be centrifuged and the clear supernatant should only be used for testing.
The use of hemolytic, lipaemic, icteric or bacterially contaminated specimens should be avoided as it may lead to erratic results.

Test procedure

Remove the test card from the foil pouch prior to use and place it on a flat and dry Surface. The test should be performed immediately after removing the test card from the foil Pouch.
Label the test cartridge with patient’s name or identification number. DO NOT write on QR Code.
Take out one D-Dimer Conjugate vial (Dried) from the aluminum pouch & immediately seal rest of the unused D-DIMER Conjugate vial (Dried) with desiccant in aluminum pouch with Clamp & rod.
Unscrew the D-Dimer Conjugate vial (Dried) & add 100 μl of Plasma sample. Close the cap of the vial by screwing action and mix well using vortexer for 30 seconds.
Keep the D-Dimer conjugate vial undisturbed for 1 minute & then vortex it well for 30 seconds.
Immediately load 50μl of above sample reagent mixture using micro-pipette to the sample well of the Cartridge. Care should be taken to avoid any spillage on the Qr Code and test result window.
Leave the sample loaded cartridge at room temperature for 20 minutes.
After 20 minutes, gently add 2 drops of assay buffer in the buffer well of the test cartridge and leave the cartridge for another 10 minutes at room temperature.
In the meantime, enter the patient’s details in the iQuant analyzer testing window and select the D-DIMER test from the pop down menu in the testing window of the iQuant analyzer.
After the incubation period is over, Insert the test cartridge into the iQuant Analyzer with arrow ( ) marked side on the top of cartridge facing towards the analyzer and press RUN button.
Note down the value displayed on the screen of iQuant Analyzer.

Limitations and interferences

The test procedure, precautions and interpretation of results for this test must be strictly followed.
As with all diagnostic tests, the test result must always be correlated with clinical finding and laboratory data available.
Any modification to the above procedure and / or use of other reagents will invalidate the test procedure.
The presence of additional substances in blood samples may interfere with product performance and may cause erroneous results.

Also, check out Urinalysis