• Categories
• Tiroides
• Órgano (Anatomía)
• Sistema endocrino
• Glándulas
• Cabeza y cuello humanos

Citation preview

Neonatal TSH Screening ELISA

ZenTech s.a. Liège Science Park Avenue du Pré Aily, 10 4031 ANGLEUR, Belgium Tel. : + 32-(0)4-361.42.32 Fax : + 32-(0)4-367.00.63 [email protected] www.zentech.be

Neonatal TSH Screening ELISA / E-GP-MZ-003 / 03-10

ENGLISH

(en)

ESPAÑOL

(es)

ISO15223

MEDICAL DEVICES SYMBOL STORAGE TEMPERATURE LIMITATION

SÍMBOLOS PARA APARATOS DE USO EN MEDICINA LIMITACION DE TEMPERATURA DE ALMACENAMIENTO

BATCH CODE

CÓDIGO DE LOTE

USE BY

CONSUMIR ANTES DE

CONSULT OPERATING INSTRUCTIONS

CONSULTAR LAS INSTRUCCIONES DE MANEJO O FUNCIONAMIENTO

IN VITRO DIAGNOSTIC DEVICE

DISPOSITIVO DE DIAGNOSTICO IN VITRO

MANUFACTURED BY

FABRICADO POR

CATALOGUE NUMBER

NUMERO DE CATALOGO

SYMBOLS (EDMA recommendations)

SÍMBOLOS (recomendaciones de la EDMA)

Number of determinations (192 - 480 - 2400)

Número de determinaciones (192 - 480 - 2400)

Calibrators

Calibradores

Control serum

Suero de control

Microtiterplate

Piaca de microvaloracion

Enzyme conjugate

HRP conjugato

Extraction solution

Solucion de Extraccion

Conjugate diluent

Diluente conjugato

Chromogen Substrate

Sustrato cromogeno

Washing solution to be diluted ten-fold

Solución de lavado que debe diluirse 10 veces

Blocking reagent

Reactivo de bloqueo

Neonatal TSH Screening ELISA / E-GP-MZ-003 / 03-10

stopped by addition of sulfuric acid and absorbance at 450 nm is read using an ELISA microtiter plate reader.

ENGLISH Enzyme immunoassay for the titration of Thyroïd Stimulating Hormone in dried blood spots of neonates. E-GP-192B (192 tests: 2x (12 strips x 8 wells)) E-GP-480B (480 tests; 5x (12 strips x 8 wells)) E-GP-2400B (2400 tests; 25x (12 strips x 8 wells))

In vitro Diagnostic (Professional use only) 1. Introduction The Neonatal TSH Screening ELISA from Zentech is an enzyme immunoassay for the quantitative determination of thyroid stimulating hormone in dried blood samples on Whatman 903 filter paper. This kit is particularly suitable for screening for primary hypothyroidism in newborns. Elevated results indicate the need for further study to assess congenital hypothyroïdism. Thyroid Stimulating Hormone (TSH) is responsible for providing the primary stimulus for the synthesis and secretion of the thyroid hormones thyroxine (T4) and triiodothyronine (T3). This glycoprotein hormone is secreted in the anterior pituitary gland, under the control of thyrotropin releasing factor (TRH), produced in the hypothalamus. The thyroid hormones produced under the direction of TSH exert a negative feedback on the pituitary gland, which regulates the secretion of TSH. This negative feedback relationship between the thyroid and pituitary glands implies that TSH is always elevated in primary hypothyroidism, often to very high levels. It is, therefore, the most sensitive test of hypothyroidism, including patients whose T4 values are still within the normal range (1, 2). Primary congenital hypothyroidism, caused by athyroidism and hypoplasia, occurs in one out of every 3,000 to 7,000 infants (3). Transient and mild hypothyroidism can also occur, especially in very preterm or very low birth weight infants (11, 13, 14). It is probably one of the most preventable causes of mental retardation. Studies have shown that the early clinical diagnosis and subsequent treatment of this disorder, usually within the first two weeks after birth, tends to prevent irreversible mental retardation, neurologic dysfunction or disabling. (4, 5,10). It has been suggested that the most effective method of assessing the infant’s thyroid function is a combination of a T4 and TSH screening program (4, 5, 6, 10, 11, 12). This is due to the fact that TSH screening may miss hypothyroidism of the secondary type, while some T4 determinations may miss minimal hypothyroidism. Therefore, the combination of T4 and TSH affords the clinician with the best possible overview of the infant’s thyroid state. Infants suspected of marginal or borderline hypothyroidism by virtue of the blood spot screening procedures should have confirmation test, performed by using serum T3, T4, and TSH determinations as well as other thyroid tests prior to initiating therapy, such as thyroid controlled stimulation by TSH, scintigraphy or sonography. Concentrations of TSH and T4 have been shown to vary with demographic variations, infant age, gender, weight, and prematurity. Therefore, it is important that each laboratory determines its own normals and cut-offs with infant age taken into account (11, 13, 14).

2. PRINCIPLE OF THE TEST

3. KIT CONTENTS 1. Coated microtiterplate: strips coated with anti TSH. 2. TSH calibrators and controls: Aluminium bag with calibrators C0C5 and internal controls L1-L2 (dried blood spots on Whatman 903 specimen collection paper using human matrix containing specified amount of human TSH). 3. TSH conjugate: 100x concentrated anti-TSH antibody conjugated to horseradish peroxidase (HRPO). Preservatives: Thimerosal (0.02%). Caution: see safety section 6. Dilute concentrated solution 1: 100 with Conjugate diluent, e.g. 300µl + 30 ml. 4. Extraction buffer: Ready for use buffer with BSA and Tween 20. Preservative: thimerosal 0.1%. 5. Conjugate diluent: Ready for use buffer with BSA and Tween 20. Preservative: thimerosal 0.1%. 6. Washing solution: 10 x concentrated Phosphate buffer with detergent. Preservative: Thimerosal 0.01%. Dilute concentrated solution 1: 10 with fresh and germ free distilled or deionised water, e.g. 55 ml + 495 ml = 550ml. Conservation of concentrated washing solution at low temperatures (2-8°C) may lead to crystal formation. These crystals dissolve upon heating (37°C) or dilution to working concentration. Stability after dilution: 10 weeks at RT (20-25° C ). 7. Chromogenic substrate: 3.3’ - 5.5’ Tetramethylbenzidine. Ready for use. 8. Blocking reagent: H2SO4, 0.5M. Ready for use. 9. Sealing tapes Quantity

Reagents Wells Calibrators 0-5 Control 1-2 TSH conjugate

Physical state

192

480

2400

2x (12x8) 1 aluminium pouch

5x (12x8) 2 aluminium pouches

25 (12x8) 4 aluminium pouches

1 x 300 µl

3 x 300 µl

13 x 300 µl

Ready for use Dried blood spots Concentrated 100x Ready for use

Extraction buffer

1 x 30 ml

3 x 30 ml

13 x 30 ml

Conjugate diluent

1 x 30 ml

3 x 30 ml

13 x 30 ml

Washing Solution

2 x 55 ml

6 x 55 ml

26 x 55 ml

1 x 30 ml

3 x 30 ml

13 x 30 ml

Ready for use

1 x 30 ml 2 sheets

3 x 30 ml 5 sheets

13 x 30 ml 25 sheets

Ready for use

Chromogenic substrate Blocking Reagent Sealing tapes

Ready for use Concentrated 10x

4. STORAGE AND STABILITY OF THE KIT When stored at 2 – 8°C unopened reagents will retai n reactivity until expiration date. Do not use reagents beyond this date. Microtiter wells must be stored at 2- 8 °C. Once th e foil pouch has been opened care should be taken to seal it tightly again.

5. MATERIAL REQUIRED BUT NOT PROVIDED

The Neonatal TSH Screening ELISA is an enzyme immunoassay to quantitate human thyroid stimulating hormone (TSH) using dried blood spots. The Neonatal TSH Screening ELISA is a sandwich ELISA. The strips are coated with an anti-TSH antibody that capture TSH present in the sample. After incubation and a washing step to remove unbound material, a monoclonal antibody anti-TSH conjugated to horseradish peroxidase (HRP) is added and allowed to bind to TSH. The following complex is formed: Mouse anti-TSH * TSH * Mouse anti-TSH-HRP conjugate After a second incubation and a washing step, the immuno-complex is detected by reduction of 3.3’-5.5’-tetramethylbenzidine (TMB) by HRP. The development of a blue colour is directly proportional to the amount of antigen in the sample or standard. The enzymatic reaction is

-

-

Single or multichannel automatic pipettes to deliver volumes in the range of 10 to 1000 µl with an accuracy of ± 1.5% over the range 10-100 µl. A microtitration plate reader capable of reading absorbances at 450 and 630 nm in dual wavelenght reading mode. A hole-punch which produces 3 mm (1/8") discs. A microtitration plate shaker (900rpm). Blood collection cards. The minimum pre-printed information required is: - infant’s name - mother’s name - patient ID number - date of birth - sex - specimen collection date

Neonatal TSH Screening ELISA / E-GP-MZ-003 / 03-10

-

submitter’s identity and address physician’s name and telephone number manufacturer and lot number of filter paper indicated on filter paper Filter paper should be Whatman903 only.

6. PRECAUTIONS A. Safety - This kit is for in vitro diagnostic only. - Washing solution and extraction buffer contain Thimerosal. This product is highly toxic by inhalation, swallowing and contact with skin. Keep away from food and drink. - Wear protective clothes and gloves. In case of contact with skin or eyes, rinse thoroughly with water. In case of accident, consult immediately a physician and show him/her the product label. - Stopping solution contains H2SO4 that is irritating for eyes and skin. Keep out of the reach of children. - Wear protective clothes and gloves. In case of contact with skin or eyes, rinse thoroughly with water and consult a doctor. - Do not smoke, eat, drink or apply cosmetics in areas where kits and blood specimens are handled. - Human blood samples should be regarded as representing a potential microbiological hazard. - Wear protective clothes and gloves when handling the samples. Samples should be incinerated after being tested. B. Operating - Do not pipette with mouth. - Do not freeze kits. - Do not use kit components after the expiration dates stated on their labels. - Do not keep diluted reagents for longer that the recommended periods (see section 3). - Keep all reagents at normal refrigerator temperature (2-8°C) in closed containers when not in use. - Ensure that all reagents are equilibrated to 18-25°C before use. - Do not use any solutions which have become turbid. - The strips, calibrators and internal controls are packed in an outer aluminium pouch containing a desiccant. Immediately after removal of strips, the remaining strips should be resealed or closed with a Scotch tape in the outer bag along with the desiccant and stored at 2-8° C. It is important to ensure the desiccant remains in the bag. - The general purpose reagents washing solution and stopping solution are interchangeable between different lots while all other reagents are specific for the individual package lot and must not be interchanged with other lots. - Do not use reagents from other manufacturers along with the kit reagents for a given test run. - Do not interchange reagent vials and their screw caps to avoid cross-contamination. Use a clean, fresh, disposable pipette tip for each reagent or specimen manipulation. - Close reagent vials tightly immediately after use to avoid evaporation and microbial contamination

7. SPECIMEN COLLECTION Blood should ideally be collected between the third and the fifth day of life (48 to 120 hours after birth). Collect blood from the infant's heel only using the medial (closest to body centre line) or lateral (furthest from body centre line) portion of the plantar (walking) surface. Blood collection from other areas of the infant foot, e.g. arch, may result in nerve, tendon or cartilage injury. Fill out the required information on the blood collection form or card (see section 5). To increase local blood flow, warm the skin puncture site using a warm (