ferritina irma ct ref

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SPECIMEN COLLECTION AND PREPARATION The assay can be performed in serum. Highly lipemic or hemolyzed samples must be discarded. Keep samples at 2-8°C for 1-2 days; for longer periods it is advisable to freeze samples at -20°C. Make sure that samples are always perfectly clear before testing. Repeated freezing and thawing of samples should be avoided. Samples with Ferritin concentrations higher than 2000 ng/mL must be diluted with the Zero Calibrator. We suggest 1:5 dilution (20 SL serum + 80 SL Zero Calibrator). ASSAY PROCEDURE − Allow reagents and samples to warm up at room temperature. − Mix samples by inversion before use. 1 - Prepare coated tubes for Calibrators and Samples. Use uncoated tubes for Total Activity. 2 - Pipette 25 DL of each Calibrator and Sample into the corresponding tubes. 3 - Pipette 500 DL of Radioactive Conjugate into all tubes. 4 - Mix the test tube rack manually. Avoid vortex mixing. 5 - Incubate for 45 minutes at room temperature on an orbital shaker (150rpm). 6 - Carefully aspirate the incubation mixture from all tubes, except those for total activity. 7 - Wash the tubes twice with 1 mL of diluted Washing Solution. Aspirate all liquid from the tubes with an aspiration pump, or decant by drying the edges of the tubes with blot-paper. 8 - Count the radioactivity in the tubes for 1 minute by using a gamma counter. We suggest to check the background of the instrument before counting the assay. In order to avoid variations in the sensitivity of the system, the background must be reduced to a minimum or adjusted properly. ASSAY SCHEME: see p. 31 CALCULATION OF RESULTS Draw a calibration curve on linear graph paper, by plotting the cpm (or B/T%) of each calibrator (y-axis) against the relative concentration (x-axis). Consider the Zero Calibrator as the first concentration of the calibration curve. Read the cpm or (B/T)% of each sample and calculate the concentration by interpolating on the calibration curve; in case of diluted samples multiply by the dilution factor. Calculate each Calibrator and Sample percent binding (B/T%), by using the following formula: Solid phase Cpm B/T % = ⎯⎯⎯⎯⎯⎯⎯⎯⎯⎯⎯ x 100 Total Activity Cpm EXAMPLE OF CALCULATION The values shown below must be considered as an example and must not be used in place of experimental data. Description cpm B/T % Ferritin Total Activity (T) 200000 Calibrator 0 ng/mL 150 0.075 Calibrator 5 ng/mL 600 0.3 Calibrator 20 ng/mL 2680 1.34 Calibrator 50 ng/mL 6000 3.0 Calibrator 250 ng/mL 22780 11.39 Calibrator 1000 ng/mL 60480 30.24 Calibrator 2000 ng/mL 80040 40.02 Sample 6800 3.4 70 ng/mL NORMAL VALUES The values reported below are indicative. We suggest that each laboratory establishes its own normal range. Men 17 - 390 ng/mL Women: - Pre-menopause 10 - 90 ng/mL - Menopause 10 - 150 ng/mL KP33CT – FERRITINA IRMA M99 – Rev.09 – 12/2004 – Pag. 10/32
PERFORMANCES OF THE ASSAY SPECIFICITY No cross-reactions were observed with the present method. SENSITIVITY The sensitivity was calculated based upon the calibration curve and expressed as the minimal dose showing a significant difference from the Zero Calibrator (mean value + 3 S.D.). This dose is 1.5 ng/mL. PRECISION Precision was evaluated upon intra- and inter-assay variability, in 3 sera at different Ferritin concentrations. Intra-assay (Repeatability) Serum Mean ± S.D. C.V. Replicates (ng/mL) % no. a 32.6 ± 1.4 4.2 10 b 144.7 ± 7.3 5.0 10 c 373.7 ± 20.2 5.4 10 Inter-assay (Reproducibility) Serum Mean ± S.D. C.V. Assays (ng/mL) % no. a 27.6 ± 2.05 7.4 10 b 160.5 ± 8.8 5.4 10 c 1120 ± 91.0 8.1 10 ACCURACY Accuracy of the method was checked by recovery and parallelism tests: Recovery Test Known amounts of Ferritin have been added to a normal serum and tested. Added Expected Measured Recovery (ng/mL) (ng/mL) (ng/mL) % S1 ---- 10 ---- S1 + 1000 1010 1025 101 S1 + 500 510 540 105 S1 + 250 260 245 94 S1 + 50 60 58 96 S1 + 20 30 31 103 Parallelism Test A high Ferritin concentration serum was tested at different dilutions with the Zero Calibrator. Dilution Expected Measured (ng/mL) (ng/mL) S2 undiluted ---- 1493 1:2 746.5 730 1:4 373.2 421 1:8 186.6 181 1:16 93.3 101 1:32 46.6 45 1:64 23.3 24.7 1:128 11.6 12.5 KP33CT – FERRITINA IRMA M99 – Rev.09 – 12/2004 – Pag. 11/32
Page 1 and 2: FERRITINA IRMA CT REF KP33CT 100 It
Page 3 and 4: DOSAGGIO IMMUNORADIOMETRICO PER LA
Page 5 and 6: − Ai sensi del D.L. italiano n. 2
Page 7 and 8: ACCURATEZZA L'accuratezza del metod
Page 9: − Orbital shaker, adjustable at 1
Page 13 and 14: IMMUN-RADIOMETRISCHER ASSAY ZUR QUA
Page 15 and 16: − Liegt kein Sicherheitsdatenblat
Page 17 and 18: 160.5 ± 8.8 5.4 10 c 1120 ± 91.0
Page 19 and 20: 6 - WASH Solution de Lavage (concen
Page 21 and 22: La lecture de la concentration des
Page 23 and 24: ENSAYO INMUNORADIOMÉTRICO PARA LA
Page 25 and 26: − La azida sódica contenida como
Page 27 and 28: a 27.6 ± 2.05 7.4 10 b 160.5 ± 8.
Page 29 and 30: RDATE Data di Riferimento / Referen
Page 31 and 32: Schema del Dosaggio, Assay Scheme,

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