【Intended use】

25OH Vitamin D total Kit (Immunofluorescence) is mainly for the in vitro quantitative measurement of 25-hydroxyvitamin D2 and D3 (25OH-D2 and 25OH-D3) in serum and plasma . It is useful as an aid in management and monitoring of regulating the concentration of calcium and phosphate in the bloodstream and promoting the healthy growth and remodeling of bone. 25OH Vitamin D might also be active in other tissues responsible for calcium transport (placenta, kidney, mammary gland …) and endocrine gland (parathyroid glands, beta cells…).

Vitamin D is the generic term used to designate Vitamin D2 and Vitamin D3. Humans naturally produce Vitamin D3 when the skin is exposed to ultraviolet sun rays. In the liver mainly, Vitamin D3 is metabolised into 25-Hydroxyvitamin D3 (25OH-D3) which is the main form of Vitamin D circulating in the body. Vitamin D3 and Vitamin D2 are also available by ingestion through food or dietary supplementation.

Vitamin D deficiency is an important risk factor for rickets, osteomalacia, senile osteoporosis, cancer and pregnancy outcomes.Vitamin D intoxication has been shown to cause kidney and tissue damages.

【Test principle】

The test uses a competitive immunodetection method . Add the sample to the sample well of the cartridge. Through chromatography, the sample reacts with the fluorescent latex particles coated with  monoclonal antibody on the conjugate pad. The complex spreads ahead along the nitrocellulose membrane captured by the covalent couple of 25OH-D3 and bovine serum albumin (BSA) on the test line. The more target material the sample contains, the less complexes the test line accumulates. The intensity of fluorescent antibody signal reflects the amount of captured 25OH-D2/D3. The immunofluorescence quantitative analyzer produced by our company can detect the concentration of 25OH-D2/D3 in the sample.

【Package】

25 Tests /Box

【Materials provided】

Contents

25   Cartridges

1    Calibration Card

25   Diluent

1    Releasing Buffer Vial

【Accessories required but not provided】

²  Immunofluorescence quantitative analyzer（NRM-FI-1000）

²  Reagent strip incubator

²  25OH Vitamin D Control

【Storage and stability】

²  Sealed: The kit must be stored at 10-30℃, valid for 18 months.

²  Opened: The cartridge must be used within 1 hour once its foil pouch is opened. Opened diluent and releasing buffer are stable for 12 months at 2-8 °C if kept capped in original container and free from contaminations.

【Warnings and precautions】

For in vitro diagnostic use.

The package insert must be carefully followed. Reliability of the test results cannot be guaranteed if there are any deviations from the package insert.

Safety precautions

CAUTION: This product requires the handling of human samples. It is recommended that all human sourced materials are considered potentially infectious and be handled in accordance with the OSHA Standard on Bloodborne Pathogens. Biosafety Level 211 or other appropriate biosafety practices should be used for materials that contain or are suspected of containing infectious agents.

Appropriate protective measures should be taken in the collection, processing, storage, mixing of the sample and testing process; once the sample and the reagent contact skin, wash with plenty of water; if skin irritation or rash occurs, get medical advice/attention.

Samples, used cartridge and disposable tips may be potentially infectious. Proper handling and disposal methods should be established by the laboratory in accordance with local regulations.

Handling precautions

Do not use the kit beyond the expiration date. The production date and expiration date are on the label.

Different batches of reagents, calibration cards and cartridges cannot be mixed.

The cartridge is disposable and cannot be reused.

Please do not use obviously damaged kit or cartridge.

Do not insert the cartridge wetted by other liquids into the analyzer, in order not to damage and pollute the instrument.

Please avoid high temperature in the lab.

The calibration card and immunofluorescence quantitative analyzer should keep away from vibratile and electromagnetic environment when used. Slight vibration of the instrument is normal. Do not pull out the cartridge during testing.

To avoid contamination, wear clean gloves when operating with reagent kits and samples.

For a detailed discussion of handling precautions during system operation, refer to the NORMAN service information.

【Specimen collection and preparation】

The serum must be collected by standard tubes or pro-coagulation tube filled inside with separation gel. The sample is advised to be tested immediately. Serum should be stored at 2-8℃; If testing is delayed more than 24 hours, then store the sample at -20℃ or below

Heat inactivated and hemolysis samples should be abandoned. If there is precipitation in the serum, centrifugation must be done before testing.

The samples must reach room temperature (10-30℃) before the test. The cryopreserved sample can only be used after complete melting, rewarming and mixing. Avoid repeating freezing and thawing samples.

【Procedure】

Preparation

Installation of the immunofluorescence quantitative analyzer（NRM-FI-1000）and incubator

Please read the operator’s manual of the immunofluorescence quantitative analyzer (NRM-FI-1000) and reagent strip incubator carefully before use.

Calibration Card installation

Each box contains a lot-specific Calibration Card to correct for lot-to-lot deviation.

Turn on the power switch, according to the system self-test prompt, waiting for the instrument to display the main interface.

Press “Assay Management”.

Insert the Calibration Card into the card slot following the direction of the arrow, then press “Read". Confirm the lot number of the Calibration Card with the reagent relevant information, then press "Ok".

Press “Return” to the main interface.

Cartridge

Cartridge needs to reach room temperature before opened.

Test procedure

1. Transfer 90μL of releasing buffer using a transfer pipette to a diluent tube.

2. Add 10 μL sample (Human serum/plasma) using a transfer pipette to the diluent tube containing releasing buffer and mix well by pipetting 10 times.

3. Leave the tube in the tube block for 5 min.

4. Remove the cartridge (after reaching room temperature), draw in 90μL of the incubated mixture  to the well and incubate it for 10 minutes in the reagent strip incubator.

5.After incubation insert the cartridge into the immunofluorescence quantitative analyzer (NRM-FI-1000) using the cartridge arrows as a guide, and click start, then the instrument will scan the cartridge automatically.

Caution

Please check the direction of the cartridge before insertion and assure the insertion is correct.

Dilution

If a sample with Vitamin D higher than the limit of the kit of 60 ng/mL and a definitive result is required, the sample should be manually diluted with normal saline and then re-assayed according to the test procedure. The maximum valid dilution ratio is 2 times, the detection limit is 120 ng/mL.

Result analysis

The analyzer automatically calculates the Vitamin D concentration in each sample by means of a calibration curve. The results are expressed in ng/mL. For further information please refer to the operator’s manual of the immunofluorescence quantitative analyzer (NRM-FI-1000).

【Explanation of results】

The test result is only for reference and it should be considered comprehensively in conjunction with clinical symptoms, health history and other laboratory test, etc. The result cannot serve as a sole factor of clinical diagnostic.

All operation must be in strict accordance with operating procedures to get the correct result. The accuracy of test results may be affected by any change to operational procedures.

【Calibration and traceability】

Calibration

Each batch of kits contains specific calibration information, stored in the Calibration Card.

Traceability

The 25OH Vitamin D total kit is traceable to an internal standard manufactured using highly purified material.

【Quality control】

The recommended control requirement for the 25OH Vitamin D total  kit is that the sample of control be tested 3 times, additional controls may be tested in conformance with local, state, and/or federal regulations or accreditation requirements and your laboratory’s quality control policy. If a control is out of its specified range, the associated test results are invalid and samples must be retested.

Each laboratory should establish its own criteria based on the following parameters:

Each new lot.

Each new shipment (even if from the same lot previously received).

Each new operator (an individual who has not run the tests for at least two weeks).

Monthly, as continued check on storage condition.

Whenever problems (storage, operator, or other) are identified

Or other times as required by your laboratory’s standard QC procedures.