Free thyroxine (FT4)

Free thyroxine (FT4) detection kit

Instructions for use (Magnetic microparticle chemiluminescence method)

Instruction of Free thyroxine（FT4）Detection Kit（Magnetic Solid Phase Chemiluminescent Immunoassay）

[Product Name]

Common name: Free thyroxine (FT4) detection kit (magnetic microparticle chemiluminescence method)

English name: Free thyroxine (FT4) Detection Kit (Magnetic Solid Phase Chemiluminescent Immunoassay)

[Packaging Specifications]

50 tests/box, 100 tests/box, 200 tests/box.

Calibrator: 6 × 0.55mL (optional), 2 × 0.55mL (optional)

Quality control product: 2 × 0.75mL (optional)

[Intended Use]

The free thyroxine (FT4) detection kit (magnetic microparticle chemiluminescence method) (hereinafter referred to as this kit) is used for in vitro quantitative detection of free thyroxine (FT4) in human serum.

Used to assist in the assessment of thyroid status, and also plays an auxiliary role in the detection and prognosis assessment of other diseases.Thyroxine T4 is secreted by the thyroid gland, with a molecular weight of 776.87. After T4 enters the blood circulation, about 99.97% binds to binding proteins, of which about 60% binds to thyroid-binding globulin (TBG), 30% binds to thyroid-binding prealbumin (TBPA), and the rest binds to albumin (Albumin). Only 0.03% of T4 is in the free state. The ratio between bound T4 and FT4 is constant. Bound T4 cannot exert its physiological function, while FT4 can interact with target cells and exert its physiological function by binding to the T4 receptor.

The normal range of FT4 is (12-22) pmol/L. In cases of hypothyroidism caused by various reasons, FT4 is significantly lower than the normal range. However, FT4 shows a normal physiological decrease after 36 weeks of pregnancy. In non-thyroid diseases, when the condition is severe, total thyroxine (TT4) decreases, but FT4 does not decrease. For pregnant women and hyperthyroid patients who often have changes in TBG, the determination of FT4 is particularly important.

Since the determination of FT4 is not affected by the serum binding protein content, serum FT4 is a sensitive indicator of in vitro thyroid function tests. Even when physiological or pathological conditions cause changes in the binding capacity and concentration of plasma thyroxine binding protein, it can still accurately reflect the function of the thyroid gland. In the early stage and follow-up of hyperthyroidism, or in cases such as low T3 syndrome caused by certain diseases, combined with FT3 and TSH determination, the diagnostic consistency can be improved.

[Test Principle]

This kit uses a competitive method: FT4 in the sample, calibrator, or quality control product competes with biotin-labeled FT4 derivative to bind to acridinium ester (AE)-labeled FT4 monoclonal antibody. Then, streptavidin-coated magnetic microparticles are added. Through the specific binding of streptavidin and biotin, the antigen-antibody complex is connected to the magnetic particles. Under the external magnetic field, the complex formed by the immune reaction is directly precipitated, and the complex is separated from other unbound substances. After washing the complex, pre-excitation solution and excitation solution are added. Under the action of pre-excitation solution and excitation solution, acridinium ester forms an unstable excited intermediate. When the excited intermediate returns to the ground state, it emits photons to form a chemiluminescence reaction, and the chemiluminescence intensity of the reaction can be detected using a chemiluminescence instrument. Within the detection range, the chemiluminescence intensity is inversely proportional to the FT4 concentration in the sample. The four-parameter Logistic equation can be used to fit and calculate the FT4 concentration in the sample.

[Main Components]

1. Product Composition

FT4 magnetic microparticle reagent: 1 bottle, streptavidin-coated magnetic microparticles;

FT4 anti-reagent 1: 1 bottle, biotin-labeled FT4 derivative conjugate;

FT4 anti-reagent 2: 1 bottle, acridinium ester (AE)-labeled FT4 antibody conjugate;

Calibrator (optional): 6 bottles or 2 bottles, respectively, containing BSA buffer with different amounts of FT4 antigen added. The calibrator value is obtained by scanning the calibrator QR code;

Quality control product (optional): 1 bottle each of high and low points, respectively, containing BSA buffer with different amounts of FT4 antigen added. The target value range of the quality control product is obtained by scanning the quality control product QR code.

Note ①: Reagents from different batch numbers of the kit are prohibited from being used interchangeably.

Note ②: Six calibration points or two calibration points can be optionally selected for the calibrator. If the calibrator and quality control product have been provided to the user, the kit does not need to be configured repeatedly.

2. Matching Reagents

The following matching reagents are not included in this product but are required for detection:

Pre-excitation solution and excitation solution: Excite acridinium ester to catalyze the generation of photons, so that the chemiluminescence intensity can be detected using a chemiluminescence immunoassay instrument.

Washing solution: Diluted for washing the reaction system.

Source: Provided by our company.

Note: In order to ensure the accuracy of the test results, the manufacturer of the matching reagents should not be changed at will during the test.

3. Calibrator Traceability

The calibrator can be traced to the enterprise's selected measurement procedure.

[Storage Conditions and Shelf Life]

The reagent is stable for 15 months when stored in the dark at 2-8℃ and should not be frozen.

After opening, the liquid reagent is stable for 30 days in the dark at 2-8℃.

【Production Date】

See product label.

【Applicable Instruments】

Cosmed SMART500S, 6500 or Tuochuang Medical TC-300 fully automated chemiluminescence immunoassay analyzer.

【Sample Requirements】

1. This kit is suitable for serum samples.

2. After blood collection, serum should be separated immediately for analysis to avoid hemolysis. Testing should be completed within 24 hours at room temperature; it is stable for 3 days at 2-8℃ and 30 days at -20℃. Avoid repeated freezing and thawing.

3. Frozen or refrigerated samples should be returned to room temperature and mixed thoroughly before use.

4. Avoid using samples with severe hemolysis or high triglyceride levels.

5. The sample volume should be no less than 200μL.

【Test Method】

Note: Please read the instructions carefully before use. Working conditions: Room temperature (15～35℃); Relative humidity ≤80%.

1. Pre-test preparation

1.1 Before the test, all reagents should be brought to room temperature, and the instrument should be preheated for at least 30 minutes;

1.2 According to the system operating instructions, load the reaction tubes (cups), add or replace the matching reagents, clean the waste liquid and tubes, and perform the liquid path filling operation.

2. Reagent and sample loading

2.1 Before loading the reagents, place the reagents to be used on a mixing device and mix thoroughly. Visually inspect that the reagent solution components should be clear, free of foreign matter, precipitates, and flocculent matter. The magnetic microparticle reagent should be a uniform suspension without obvious aggregation;

2.2 According to the system operating instructions, scan the reagent barcodes to complete the loading of the magnetic microparticle reagent and the antibody reagent;

2.3 After mixing the calibrators and quality control samples, transfer them to the instrument's dedicated reaction cups, load them into the instrument's sample positions, and load the centrifuged serum samples directly into the instrument's sample positions after scanning.

3. Detection steps

For optimal detection performance, please follow the relevant guidance in this instruction manual and operate according to the operating manual of the fully automated chemiluminescence analyzer.

Note: The detection reaction process and related parameters have been predefined in the instrument operating software.

4. Calibration curve and calibration

The calibration curve is obtained by scanning the main curve card to generate it directly, or by directly detecting six calibrator points to generate a fit, and the system also supports a two-point calibration method to obtain the working calibration curve.

Note ①: Due to inter-instrument differences and system differences caused by different operators, operating environments, and matching general reagents, the six-point direct calibration method is recommended to obtain the working calibration curve.

Note ②: Due to reagent activity drift and changes in batches of general reagents, the calibrated working curve needs to be recalibrated after a certain period of use. Recalibration should be performed under the following circumstances:

l After 1 month (28 days) of using the same batch number of reagents;

l When using a new batch of reagents, the whole box or pre-activation liquid and activation liquid;

l When the quality control values are outside the quality control range;

l After each maintenance of the analyzer.

5. Result output

The analyzer automatically calculates the FT4 concentration of each sample using a working curve obtained by two-point calibration from the standard curve, and the results are expressed in pmol/L.

6. Quality control

Two levels of quality control samples should be measured daily along with sample testing, treating the quality control samples as patient samples;

Note: It is recommended to use at least two levels of commercial quality control products. If the measured results are within the system's acceptable quality control range or within your specified range (determined by an appropriate internal laboratory quality control program), the test results are satisfactory; otherwise, it indicates that the test results are unreliable and a test report should not be issued.

【Reference Range】

Normal range of this kit: 12.0-22.0 pmol/L

Note: The FT4 levels measured will vary depending on the region, individual, and method used. Therefore, we recommend that each laboratory establish its own normal range. A diagnosis should not be made based solely on the FT4 value obtained by this method; the results should be analyzed in conjunction with other clinical data, including the patient's specific condition and treatment status.

【Interpretation of Test Results】

1. Due to methodological or antibody specificity reasons, testing the same sample using reagents from different manufacturers may yield different results. Therefore, results obtained using different reagents should not be directly compared to avoid erroneous medical interpretation.

2. Test results exceeding the measurement range of the kit are calculated results extrapolated from the calibrator curve. When reporting such results, please pay special attention. If an accurate value is desired, dilute the sample appropriately before testing; the maximum dilution factor is 10 times.

3. Quality control products can serve as a reference for the reliability of the results of the current experiment; their measured values should be within the allowable range specified on the quality control sheet for this batch of products. Test results should be judged comprehensively based on the reference range and other clinical factors and results. When the test result is close to the upper limit of the reference range, consider performing a confirmatory test on the sample.

【Limitations of the Test Method】

1. Severe hemolysis (hemoglobin ≥500mg/dL), lipemia (triglycerides ≥1500mg/dL), jaundice (bilirubin ≥20mg/dL), and contaminated samples may affect the test results and should be avoided.

2. The test results of this kit are for clinical reference only and cannot be used alone as the basis for confirming or excluding cases. To achieve diagnostic purposes, this test result should be used in conjunction with clinical examinations, medical history, and other examinations.

3. This product can be used for the determination of human serum samples. The reliability of determining FT4 concentration in other body fluid samples has not been fully confirmed.

Product Performance Indicators

1. Reagent Performance Indicators

1.1 Accuracy: The recovery rate P should be within the range of 85%～115%.

1.2 Precision: Intra-batch precision ≤8%, inter-batch precision ≤10%.

1.3 Blank limit: Should not be greater than 3 pmol/L.

1.4 Linearity: Within the range of (3～100) pmol/L, the linear correlation coefficient r should be ≥0.9900.

1.5 Specificity: When measuring samples containing 500 ng/mL of TT3 and 50 ng/mL of rT3, respectively, the measured results should not be higher than 3 pmol/L.

2. Calibrator Performance Indicators

2.1 Accuracy: The recovery rate P should be within the range of 85%～115%.

2.2 Homogeneity: Inter-bottle difference (CV) ≤8%.

3. Quality Control Product Performance Indicators

3.1 Expected results: The test value should be within ±15% of the target value of the quality control product.

3.2 Homogeneity: Inter-bottle difference (CV) ≤8%.

【Precautions】

1. This reagent is for in vitro diagnostic use only. Reagents from different batches are prohibited from being used interchangeably. Do not use expired reagents.

2. Some components of this kit contain bovine-derived materials, but these raw materials are not from countries or regions with bovine spongiform encephalopathy (BSE) epidemics, and their production is not carried out in countries or regions with BSE epidemics. If it is bovine serum, it will be filtered and sterilized after purchase before being used in kit production. Therefore, the kit does not pose a risk of transmitting BSE; some components of the kit contain porcine-derived materials, but these raw materials are not from countries or regions with foot-and-mouth disease epidemics, and their production is not carried out in countries or regions with foot-and-mouth disease epidemics. If it is porcine serum, it will be filtered and sterilized after purchase before being used in kit production. Therefore, the kit does not pose a risk of transmitting foot-and-mouth disease; this reagent does not contain human serum, but it should still be regarded as a potential biohazard.

3. This reagent contains preservatives. Avoid contact with skin. If skin accidentally comes into contact with the reagent, rinse thoroughly with plenty of water.

4. All reagents should be stored at 2～8℃, avoid freezing at -20℃, and equilibrate to room temperature before use.

5. The magnetic microparticle reagent should be thoroughly mixed before use, and strong shaking should be avoided.

【Explanation of Labels】

Store at 2-8℃ Avoid direct sunlight Store upright

In vitro diagnostic reagent Production date Expiration date

  Product batch number See instructions for details

【References】

1. Zhang, Y., & Yang, J. (2009). Correlation analysis of serum free thyroxine and dyslipidemia in individuals with normal thyroid function. Shanxi Medical Journal, 38(z1), 12.

2. Oppenheimer JH. Role of plasma proteins in the binding, distribution and metabolism of the thyroid hormones. N Engl J Med. 1986,278:1153-1162.

3. Tabachnick M, Giorgio NA Jr. Thyroxine Protein Interactions Arch Biochem Biophys 1964; 105:563-569.

4. Hou, Z., & Wang, F. (2016). Advances in immunological detection methods for thyroid hormones. Medical Review, 22(02), 130-133.

【Basic Information】

Registrant/Manufacturer: Zhejiang Tuochuang Medical Technology Co., Ltd

Address: No. 1 Building, Qianren Plan Industrial Park, No. 78, Zhancheng Avenue, Zhuji City, Zhejiang Province

Production Address: No. 1 Building, Qianren Plan Industrial Park, No. 78, Zhancheng Avenue, Zhuji City, Zhejiang Province

After-sales service unit: Zhejiang Tuochuang Medical Technology Co., Ltd

Production license number:

Contact number: 0575-87178692

Fax:

Post code: 311800

Website: www.zjtcyl.com

【Medical Device Registration Certificate Number/Product Technical Requirements Number】

【Date of Approval and Revision of Instructions for Use】

Approval date:

Revision date: